Review Article

Apolipoproteins: metabolic role and clinical biochemistry

applications

Marek H Dominiczak1 and Muriel J Caslake2

1
NHS Greater Glasgow and Clyde Clinical Biochemistry Service and College of Medical, Veterinary and Life Sciences, University of
Glasgow, Department of Biochemistry, Gartnavel General Hospital, 1053 Great Western Road, Glasgow G12 0YN; 2Vascular Biochemistry
Section, Institute of Cardiovascular & Medical Sciences, College of Medical, Veterinary & Life Sciences, University of Glasgow, 2nd floor
McGregor Building, Western Infirmary, Glasgow G11 6NT, UK
Corresponding author: Marek H Dominiczak. Email: marek.dominiczak@gla.ac.uk

Abstract
Lipoprotein metabolism is dependent on apolipoproteins, multifunctional proteins that serve as templates for the assembly of
lipoprotein particles, maintain their structure and direct their metabolism through binding to membrane receptors and
regulation of enzyme activity. The three principal functions of lipoproteins are contribution to interorgan fuel (triglyceride)
distribution (by means of the fuel transport pathway), to the maintenance of the extracellular cholesterol pool (by means of the
overflow pathway) and reverse cholesterol transport. The most important clinical application of apolipoprotein measurements
in the plasma is in the assessment of cardiovascular risk. Concentrations of apolipoprotein B and apolipoprotein AI (and their
ratio) seem to be better markers of cardiovascular risk than conventional markers such as total cholesterol and LDL-
cholesterol. Apolipoprotein measurements are also better standardized than the conventional tests. We suggest that
measurements of apolipoprotein AI and apolipoprotein B are included as a part of the specialist lipid profile. We also suggest
that lipoprotein (a) should be measured as part of the initial assessment of dyslipidaemias because of its consistent
association with cardiovascular risk. Genotyping of apolipoprotein E isoforms remains useful in the investigation of mixed
dyslipidaemias. Lastly, the role of postprandial metabolism is increasingly recognized in the context of atherogenesis, obesity
and diabetes. This requires better markers of chylomicrons, very-low-density lipoproteins and remnant particles.
Measurements of apolipoprotein B48 and remnant lipoprotein cholesterol are currently the key tests in this emerging field.

Ann Clin Biochem 2011; 48: 498 515. DOI: 10.1258/acb.2011.011111

Introduction Lipoprotein metabolism

Abnormalities in lipoprotein metabolism are important in
atherogenesis, obesity, insulin resistance and diabetes; all
being major areas of concern for public health, individual
wellbeing and research. Control of lipoprotein metabolism
through lifestyle measures and medication has become an
essential part of cardiovascular prevention.
This paper aims to provide a perspective on apolipopro-
teins and their measurement in the context of lipid metab-
olism and associated clinical disorders.
First, lipid metabolism is briey reviewed, with an empha-
sis on the role of apolipoproteins as markers of lipoprotein
particles and as metabolic agents. The apolipoproteins are
then discussed under the three headings: structure and func-
tion, clinical signicance and methodology.
This paper was prepared at the invitation of the Clinical Sciences
Reviews Committee of the Association for Clinical Biochemistry.
Lipoproteins are lipidated protein particles that carry
hydrophobic substances in the hydrophilic environment of
plasma. They are classied on the basis of their hydrated
density in an ascending order into chylomicrons,
very-low-density lipoproteins (VLDL), intermediate-density
lipoproteins (IDL), low-density lipoproteins (LDL) and
high-density lipoproteins (HDL). The term IDL is often
used interchangeably with remnant particles. However,
because the VLDL remnants may distribute both in the
IDL fraction (1.006 1019 kg/L) and the VLDL fraction
(,1.006 kg/L), the equivalence is not complete. For a
general outline of lipid metabolism, the reader is referred
to a current textbook,1 and literature on earlier lipid research
can be found in a number of previous reviews.2 6
The surface of a lipoprotein particle consists of either
hydrophilic or amphipathic substances such as phospholi-
pids and cholesterol as well as apolipoproteins, which

Annals of Clinical Biochemistry 2011; 48: 498 515

 Dominiczak and Caslake. Apolipoproteins: theory and applications 499
................................................................................................................................................

serve as receptor-binding and regulatory proteins. The cores
of these particles contain hydrophobic molecules such as tri-
glycerides (TG) and cholesteryl esters.
Lipoproteins differ in size, lipid composition and apolipo-
protein content. These characteristics change as a result
of action of enzymes such as lipoprotein lipase (LPL),7
hepatic triglyceride lipase (HTGL),8 lecithin-cholesterol
acyltransferase (LCAT)9,10 and cholesteryl ester transfer
protein (CETP).11 Apolipoproteins regulate the activities of
these enzymes, and can transfer between different lipopro-
tein classes. Lipoprotein assembly and metabolism are
affected both by apolipoprotein content and by their confor-
mation. Apolipoproteins control cellular uptake of lipopro-
teins through binding to membrane lipoprotein receptors.
Apolipoprotein B100 (apoB100) and apolipoprotein E
(apoE) bind to the apoB/E (LDL) receptor.12
Apolipoprotein E (apoE) also binds to the LDL receptor-
related protein (LRP).13 Apolipoproteins A bind to the sca-
venger receptor BI.14
Lipoproteins contribute to body fuel metabolism by
enabling TG distribution between tissues. They also serve
as an extracellular reservoir, and a vehicle for transport, of
cholesterol. Plasma TG measurements reect the rst,
and measurements of total cholesterol (TC) and
LDL-cholesterol (LDL-C) the second and third function.
Clinical disorders may result from both defects in fuel trans-
port and in cholesterol homeostasis. Often a defect in one
process will secondarily affect the other.
Abnormalities in fuel metabolism are present in cardio-
vascular disease (CVD), obesity and diabetes mellitus
conditions representing the three major contemporary
epidemics. Hypercholesterolaemia is related to the entry of
cholesterol-containing lipoprotein particles into vascular
intima and thus atherogenesis,15 while hypertriglyceridae-
mia, apart from its association with cardiovascular
disease, leads to steatosis and may precipitate pancreatitis.
Discussion of atherogenesis is beyond the scope of this
article but it has been extensively reviewed elsewhere.16,17
Pathways of lipoprotein metabolism
To describe lipoprotein metabolism, this paper employs the
conceptual framework of the fuel transport pathway
(metabolism of chylomicrons, VLDL and remnant particles),
the overow pathway (the metabolism of LDL) and reverse
cholesterol transport (the metabolism of HDL).18 We believe
that this approach facilitates understanding of the

HDL

E C
B48 Lipoprotein
AV lipase
AI AII AIV B48 E
AI AII AIV AV
Nascent CI CII, CIII
chylomicrons
Mature
AIV chylomicrons
PERIPHERAL TISSUES

CE
INTESTINE Chylomicron
B48 HDL
remnants
EC
AI AII AV C
TG
B48
C
E B48
C
E
LDL receptor

LRP

ARTERY

LIVER

Figure 1 Apolipoproteins in the fuel transport pathway of lipoprotein metabolism: the chylomicrons. Chylomicrons transport triglycerides from the intestine to
the peripheral tissues. Apolipoprotein B48 (present in the concentration of one molecule per particle) tracks a particle from a nascent chylomicron to a chylo-
micron remnant and is the most important marker of chylomicron metabolism. Note extensive apolipoprotein exchanges between different particles. The
measurement of apolipoprotein B48 reflects both nascent chylomicrons and chylomicron remnants. On the other hand, the measurement of remnant lipoprotein
cholesterol (RLP-C) reflects the sum of chylomicron remnants and VLDL remnants (compare Figure 2) but excludes nascent or mature chylomicrons. AI, apoli-
poprotein AI; AII, apolipoprotein AII; AIV, apolipoprotein AIV; AV, apolipoprotein AV; B48, apolipoprotein B48; CI, apolipoprotein CI; CII, apolipoprotein CII; CIII,
apolipoprotein CIII; E, apolipoprotein E; CE, cholesteryl esters; LRP, LDL receptor-like protein; TG, triglycerides
500 Annals of Clinical Biochemistry Volume 48 November 2011
................................................................................................................................................

pathogenesis of lipid disorders and relevant therapeutic
interventions.
The fuel transport pathway
The fuel transport pathway delivers TG to peripheral
tissues. The TG transport function is shared by chylomi-
crons and VLDL. While chylomicrons are assembled
during the absorptive phase of the feed-fast cycle, VLDL
can be present in plasma at any time.
TG are incorporated into chylomicrons in the enterocytes,
on the template of apoB48 and are secreted into the lymph
(Figure 1).19 Nascent chylomicrons contain apoB48, apolipo-
protein AIV (apoAIV) and apolipoprotein AV (apoAV).
Later, they acquire ( probably from HDL) apoAI and
apoAII and apolipoproteins C (apoCI, CII, CIII) as well as
apoE. ApoAIV is released into plasma.
In the periphery, chylomicron TG are digested by LPL, an
enzyme present in the vascular endothelium. Fatty acids are
released to cells. Chylomicrons decrease in size and become
chylomicron remnants. Now apoA, most apoC, and prob-
ably apoAV, are returned to the HDL. The remnants are
internalized in the liver after binding to the LRP and to
the LDL receptor, mediated by apoE.
In contrast to TG absorbed from the intestine, endogenous
TG are secreted from the liver in VLDL particles (Figure 2).
Nascent VLDL contain apoB100 as a sole apolipoprotein.
They subsequently acquire (mostly from HDL) apoA (AI,
AII, AIV), apoC (CI, CII and CIII) and apoE. VLDL TG
are, analogously to the chylomicrons, partially digested by
LPL. This transforms VLDL particles into VLDL rem-
nants.20,21 VLDL remnants are internalized through the
LDL receptor, to which they bind through apoE but not
apoB. Note that the TG-rich particles in the fuel transport
pathway lose some TG to HDL, in exchange for cholesteryl
esters,22 in a process mediated by the CETP.23,24
The overflow pathway
Internalization of the remnants returns these particles to the
liver. This would neatly complete the fuel transport
pathway, if not for the fact that the internalization occurs
in parallel to further lipolysis, this time by the HTGL,
which is bound to endothelium in the hepatic microvascula-
ture. During this process, the remnants lose apoC, and apoE
(which are transferred to HDL), decrease in size, and
become LDL.25 Thus the LDL particles are generated as a
result of overow from the fuel transport pathway. This
process is clinically important, because when remnant
supply from the fuel transport pathway is excessive due
to oversupply or inefcient removal more LDL particles
are generated.
The overow pathway involves further metabolism of
LDL (Figure 3). ApoB100, the main apolipoprotein of the
LDL, controls their internalization by binding to the
apoB/E (LDL) receptor. It has lesser afnity to the LDL
receptor than apoE in the remnants, and the LDL residence
time in the circulation exceeds that of the remnants.

CE
Lipoprotein lipase
HDL B100
B100
E C E
TG AI AII AIV
CI CII CIII

PERIPHERAL TISSUES

HDL
CE
VLDL B100
E
EC remnants HDL
LDL AI AII AIV
B100 TG

B100
VLDL HTGL EC

B100 LDL receptor

AI AII AIVI

Nascent VLDL
LIVER ARTERY

Figure 2 Apolipoproteins in the fuel transport pathway of lipoprotein metabolism: the VLDL. Apolipoprotein B100 tracks the metabolism of VLDL. It is present in
the concentration of one molecule per particle. Note the parallels with chylomicron metabolism (compare Figure 1) in particular with regard to apolipoprotein
exchanges with HDL. The fuel transport pathway generates LDL through the action of HTGL. The measurement of apoB100 includes all the particles in this
pathway together with all LDL particles in the overflow pathway (compare Figure 3). AI, apolipoprotein AI; AII, apolipoprotein AII; AIV, apolipoprotein AIV; AV, apo-
lipoprotein AV; B100, apolipoprotein B100; CI, apolipoprotein CI; CII, apolipoprotein CII; CIII, apolipoprotein CIII; E, apolipoprotein E; CE, cholesteryl esters;
HTGL, hepatic triglyceride lipase
 Dominiczak and Caslake. Apolipoproteins: theory and applications 501
................................................................................................................................................

HDL
PERIPHERAL TISSUES

B100
EC VLDL EC
B100 remnants

Range of sizes:
buoyant and
small dense LDL B100 B100
LDL
HTGL

LDL ARTERY
receptor

PERIPHERAL
TISSUES
B100
B100
LIVER

LDL receptor

Figure 3 Apolipoproteins in the overflow pathway of lipoprotein metabolism: the LDL. The name overflow pathway reflects the fact that LDL are generated from
the fuel transport pathway. The oversupply of remnants would increase the number of generated LDL. The predominant lipoprotein of LDL is B100, present in the
concentration of one molecule per particle. The measurement of apoprotein B100 reflects the sum of LDL particles, the VLDL and the VLDL remnants. Note that
LDL is generated in a range of sizes. The main determinant of their plasma concentration is the activity of the LDL receptor and the rate of cellular uptake. B100,
apoprotein B100; HTGL, hepatic triglyceride lipase. Note: the empty circles in the upper part of the figure represent lipoproteins in the fuel transport pathway (refer
to Figures 1 and 2)

The LDL serve as an extracellular cholesterol depot: their
cellular uptake increases when LDL receptor expression
increases as a result of a decrease in the intracellular choles-
terol concentration.
Reverse cholesterol transport and cholesterol recycling
Metabolism of HDL, known as reverse cholesterol transport
(RCT), is a pathway of removal of cholesterol from periph-
eral cells to the liver (Figure 4).26 28 This view is now
being challenged, since HDL also remove cholesterol from
the liver inserting it back into the VLDL and remnant par-
ticles,26 through the exchange mechanism mentioned
above. Transport of cholesterol from macrophages to the
liver (macrophage RCT: the central concept in atherogen-
esis) involves only a small fraction of circulating HDL. It
seems therefore that HDL, similarly to LDL, contribute to
the maintenance of the extracellular cholesterol pool.
HDL are assembled on the matrix of the apoA. ApoA is
secreted by the liver and intestine and is lipidated extracel-
lularly, transforming into nascent ( pre-beta) HDL,
phospholipid-rich, discoid particles. They acquire choles-
terol by binding to the membrane ATP-binding cassette
transporter A1 (ABCA1) that controls efux of free choles-
terol from cells. Cholesterol is subsequently esteried by
LCAT. Accumulation of cholesteryl esters changes the
particle to spherical HDL3. HDL3 transfer cholesteryl
esters, apoA, apoC and apoE to TG-rich lipoproteins in
exchange for TG (see above). This process enlarges the
HDL particles which became HDL2. HDL2 subsequently
ofoad cholesteryl esters by docking into the scavenger
receptor BI (there is no particle internalization).
Conventional markers of lipoprotein
metabolism
The conventional routine markers of lipoprotein metab-
olism include TC, LDL-cholesterol (LDL-C), HDL-choles-
terol (HDL-C) and plasma TG. More detailed assessment
of a lipid prole includes ultracentrifugation-based
analysis. Since lipoprotein components such as cholesterol
or TG are present in more that one lipoprotein class,
neither of them can serve as an absolute marker of a parti-
cular class. Moreover, LDL, similarly to VLDL, are hetero-
geneous in size.29,30 All this means that measurement of
LDL-C is not a good indicator of the number of LDL
particles.
The most commonly measured substance, cholesterol, is
measured either in the plasma (TC)31 or in lipoprotein frac-
tions (LDL-C, HDL-C). TC predominantly reects the con-
centration of LDL particles. HDL-C constitutes around
25% of TC. VLDL normally constitute only approximately
502 Annals of Clinical Biochemistry Volume 48 November 2011
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Lipid-poor
AI AII E
Intestine
PERIPHERAL TISSUES

Lipid-poor
ApoAI
Cholesterol
ABCA1
HDL2

CE
AI AII
Nascent
(pre-beta) HDL Scavenger
receptor B1
LCAT
alpha- HDL
ARTERY

HDL3
LIVER
CE
AI AII C
TG

Chylomicrons HDL2
VLDL
Remnants

Figure 4 Apolipoproteins in reverse cholesterol transport: the HDL. HDL particles transport cholesterol from peripheral cells and macrophages (macrophage
RCT) to the liver but also insert cholesteryl esters into the fuel transport pathway by exchanging them for TG with TG-rich proteins. In this way, HDL contributes
to the maintenance of the extracellular cholesterol pool. The measurement of ApoAI is the most important marker of the activity of reverse cholesterol transport.
AI, apolipoprotein AI; AII, apolipoprotein AII; AIV, apolipoprotein AIV; AV, apolipoprotein AV; CI, apolipoprotein CI; CII, apolipoprotein CII; CIII, apolipoprotein CIII;
E, apolipoprotein E; CE, cholesteryl esters; B48, apolipoprotein B48; LCAT, lecithin cholesteryl-ester transferase; CE, cholesteryl-esters; ABCA1, ATP-binding
cassette transporter A1; RCT, reverse cholesterol transport; TG, triglycerides

15% of the total. Measuring HDL-C together with TC
enables computing the TC/HDL-C ratio, assumed to
reect the balance between transport of cholesterol to the
periphery and its return to the liver.
The measurement of TG, the third component of the tra-
ditional fasting lipid prole, reects the concentration of
VLDL and remnant particles, and, postprandially, chylomi-
crons and their remnants. In routine practice, measurements
of TC, HDL-C and TG allow calculation of LDL-C by the
Friedewald formula:32
LDL-C mmol=L TC  HDL-C  (TG=2:2)
Thus LDL-C is estimated by taking into account the
HDL-C and TG concentration. LDL-C calculated in this
way in fact includes VLDL-C, IDL-cholesterol (IDL-C) and
lipoprotein (a) (Lp(a))-cholesterol. Naturally, this calculation
incorporates the errors of TC, HDL-C and TG measure-
ments. The formula is not applicable at TG concentrations
above 4.5 mmol/L. In spite of these limitations, the
formula has been commonly used for the assessment of car-
diovascular risk and particularly for the monitoring of
lipid-lowering treatment.
More recent is the calculation of non-HDL cholesterol
(NHDL-C). This is calculated as:
NHDL-C TC  HDL-C
NHDL-C has been recommended as superior to the
LDL-C. NHDL-C is a simple estimate of a sum of cholesterol
contained in VLDL, remnant particles, LDL and Lp(a). It
seems to be a better predictor of coronary heart disease
(CHD) than LDL-C.33 36 Its use has been recommended
by the American Diabetes Association and the American
College of Cardiology.
The gold standard method of measuring lipids and
lipoproteins (the so-called beta-quantication) employs
ultracentrifugation to partially separate lipoproteins.37
After removal of VLDL by ultracentrifugation, the other
apoB-containing lipoproteins in the infranatant are precipi-
tated using heparin and manganese chloride, leaving HDL
in solution for cholesterol measurement. LDL-C is calcu-
lated as:
LDLC TC  VLDL-C  HDL-C
Note that here the calculated LDL-C value also includes
IDL-C and Lp(a) cholesterol. This method is available in
the UK in specialized lipid laboratories.
 Dominiczak and Caslake. Apolipoproteins: theory and applications
................................................................................................................................................
As already mentioned, the LDL-C measurement reects
just one component of LDL particles and cannot provide
precise estimation of the LDL particle number, an important
parameter relating to LDL atherogenecity.
In Friedewalds paper, the differences between calcu-
lated and measured LDL, after exclusion of patients with
TG above 4.5 mmol/L (400 mg/dL), were 4.8 9.8%
depending on the type of dyslipidaemia.32 When
Friedewald-formula-derived LDL-C and LDL-C obtained
through beta-quantication were compared across ve cat-
egories of LDL-C concentration, the misclassication rates
(instances where a result obtained by each method would
fall into a different category) were 20 30%. On the other
hand, using standardized methodology of apolipoprotein
measurement and reference material SP-07, the between-
laboratory coefcient of variation for apoA was 2.1 5.6%
and for apoB 3.1 6.7%.38
Thus there are both technical inadequacies and scientic
limitations inherent in conventional methods of lipid and
lipoproteins assessment. For a more detailed account of
the conventional tests, the reader is referred to other
reviews.31,39
Structure and function of apolipoproteins
Please refer to Table 1 throughout this section.
Apolipoprotein B100
ApoB100 consists of 4536 amino acids and has a molecular
mass of 550,000 Da. After cleavage of 27 amino acids from
the N-terminal end, the molecular mass decreases to
513,000 Da.40,41 It possesses a globular amino-terminal
domain, which reacts with microsomal TG transfer pro-
teins.42 The gene for apoB is located on the short arm of
chromosome 2. One hundred and twenty-three genetic var-
iants in the apoB gene were identied in the Copenhagen
City Heart Study. ApoB polymorphisms include T2488T
and E4154K,43 which are signal peptide insertion/deletion
polymorphisms. Mutation of apoB100 at amino acid
residue 3500 affects its receptor-binding and leads to a dis-
order known as familial defective ApoB (FDB).44 ApoB100
is synthesized in the hepatocytes and its excess is degraded
within cells. It resides in the VLDL, LDL and LDL particles
at a ratio of one molecule per lipoprotein particle. Thus, it is
a marker of the number of these particles.
Apolipoprotein B48
ApoB48 has 2152 amino acids and a molecular mass of
264,000 Da. It is a truncated form (amino-terminal 40%) of
apoB100.45 ApoB100 and apoB48 are produced from a
single gene. During the editing process of the apoB
mRNA, deamination of a cytidine nucleotide 6666 to
uridine converts that codon to a stop codon. This results
in synthesis of apoB48. ApoB48 is synthesized by entero-
cytes and secreted in chylomicrons, and is retained in chylo-
micron remnants. Since one molecule of apoB48 is present in
each chylomicron particle, it serves as a marker of the
503
number of chylomicrons and their remnants. ApoB48 does
not possess the LDL-receptor binding domain: the remnants
are internalized by the apoE-binding LRP and also to a
degree by the LDL receptor, to which they also bind
through apoE.46
Particles smaller than 70 80 nm can penetrate the vascu-
lar wall. Therefore, chylomicron remnants, but not nascent
chylomicrons, are regarded as atherogenic. Chylomicron
remnants are enriched in cholesteryl esters through
exchange with HDL (see above).47
Apolipoprotein AI
ApoAI is a protein of 243 amino acids. The apoA gene is
located on chromosome 11, and is part of the APOA1/C3/
A4/A5 gene cluster.48 It is synthesized in the liver and intes-
tine.49 The concentration of apoAI is controlled by its degra-
dation rate: apoAI that cannot acquire lipids is rapidly
degraded. ApoAI constitutes 70% of HDL apolipoproteins.
It activates LCAT and is also an anti-inammatory molecule
and an antioxidant.50 Plasma levels of apoAI vary and
depend on method of measurement and population.
Apolipoprotein AII
ApoAII is a 77-amino acid homodimer with molecular mass
of 17,400 Da. It is predominantly expressed in the liver.
ApoAII accounts for approximately 20% of HDL protein.
HDL particles contain just apoAI (these are designated as
LpAI) or both apoAI and apoAII (LpAI/AII).51 ApoAII con-
centration is determined by its production rate. It inhibits
LPL activity (and may also inhibit HTGL), and serves as a
co-factor for LCAT and CETP. It may replace apoCII (a
LPL activator) in the VLDL, or directly inhibit the LPL.
LPL activation by the HDL particles was impaired in trans-
genic mice expressing human apoAII.52 Experiments in
transgenic mice also demonstrated that apoAII is associated
with obesity and insulin resistance. In contrast to the results
of mouse studies,53 it seems that it may be inversely associ-
ated with the risk of coronary disease in humans.54 ApoAII
has been reviewed by Tailleux et al. 55
Apolipoprotein AIV
ApoAIV is a glycoprotein with molecular weight of
46,000 Da. It is synthesized in the intestine and is incorpor-
ated into nascent chylomicrons. It is also present in HDL.
Because it is relatively hydrophilic, it may be displaced
from lipoproteins by apoCII and apoE and circulates predo-
minantly as lipid-free protein. It participates in intestinal
lipid absorption, in the assembly of chylomicrons and
affects various aspects of lipoprotein response to diet. Its
polymorphism Q360H leads to an increased postprandial
TG concentration, a reduced LDL response to dietary
cholesterol and an increased HDL response to dietary fat.
The effects of T347S polymorphism are opposite to
Q360H. ApoAIV also modulates the activity of LPL and
activates LCAT.56
504 Annals of Clinical Biochemistry Volume 48 November 2011
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Table 1 Structure and function of apolipoproteins

Lipoprotein
Examples of metabolism
Apo Genes isoforms Synthesis Structure Function Lipoproteins pathway
AI Chromosome 11, Six polymorphic Liver, 243 AA, Structural in HDL LCAT 70% of HDL RCT, fuel
AI/C3/A4/A5 isoforms. intestine 28,000 Da activator protein. Most transport
gene cluster Mutations: Apo abundant pathway
AI Tangier, AI protein in HDL.
Milano AI Chylomicrons,
Marburg VLDL
AII Chromosome1 Liver, 77AA, 17,400 Da. Structural in HDL TG and 20% of HDL RCT (main
intestine Mainly present fatty acid metabolism. protein. Second marker), fuel
as dimer (mol. Links with apoE in HDL most abundant transport
mass above is after apoAI. pathway
that of the Chylomicrons,
dimer) VLDL
AIV Chromosome 11, ApoAIV 360 Liver, Metabolism of TG-rich Chylomicrons, Fuel transport
A1/C3/A4/A5 (common), intestine particles. Interacts with HDL, free in pathway, RCT
gene cluster ApoAIV-1, CII in LPL. LCAT activator plasma
ApoAIV-2
AV Chromosome 11, Multiple variants Liver Chylomicron- and VLDL Chylomicrons, Fuel transport
A1/C3 A4/A5 assembly. LPL activator VLDL, HDL pathway, RCT
gene cluster
CIII Chromosome 11, Variants with Liver, 79AA, 8,800 Da LPL inhibitor. Masks or Surface of TG-rich Fuel transport
A1/C3 A4/A5 differing sialic intestine displaces apoE from LRP particles: pathway, RCT
gene cluster acid content: chylomicrons,
CIII-0, CIII-1, VLDL, remnants
CIII-2 HDL
CII Chromosome 19 Liver, 79AA, 8900 Da LPL activator: deficiency Chylomicrons, Fuel transport
intestine leads to gross VLDL, HDL pathway, RCT
hypertriglyceridaemia
CI Chromosome 19 Liver, 57AA LCAT activator, LPL Chylomicrons, Fuel transport
intestine inhibitor, CETP inhibitor. VLDL, HDL pathway, RCT
Inhibits apoE binding to
LRP
B100 Chromosome 2 Over 100 Liver 4536 AA, Structural component of VLDL, IDL, LDL Fuel transport
polymorphisms 550,000 Da VLDL, IDL, LDL. Ligand pathway,
for LDL-receptor overflow
pathway. One
molecule per
particle
marker of
particle
number
B48 Chromosome 2 Intestine 2152 N-terminal Structural component of Chylomicrons, Fuel transport
AA of B100, chylomicrons and chylomicron pathway
264,000 Da. chylomicron remnants remnants
8 10% CHO
E Chromosome 19, Three main Liver, 299 AA, Multifunction protein. Chylomicrons, Fuel transport
E/C1/C2/C4 isoforms: e2, intestine, 34,200 Da LDL-receptor ligand for VLDL, remnants pathway, RCT
gene cluster e3, e4. Many brain, LDL and chylomicron HDL
variants kidney, remnants. LRP ligand.
spleen, Role in RCT. Modulates
adrenals LPL, CETP, LCAT, HTGL.
and other Antioxidant molecule.
Regulator of inflammatory
response
(a) Chromosome Over 20 isoforms, Liver Variable HDL2, LDL Role in
6. Linkage with dependent on molecular fibrinolysis?
plasminogen number of mass: 187,000
gene. Kringle 4 kringle 4 800,000 Da
region most repeats Pre-beta
variable mobility. High
sialic acid
content

CHO, carbohydrates; AA, aminoacids; RCT, reverse cholesterol transport; LRP, LDL receptor-like protein
Please see text for references
 Dominiczak and Caslake. Apolipoproteins: theory and applications
................................................................................................................................................
Apolipoprotein AV
ApoAV modulates hepatic VLDL synthesis and secretion.
The gene coding for apoAV (APOA5) has been strongly
associated with TG concentration. ApoAV deciency leads
to type V dyslipidaemia and to reduced post-heparin LPL
activity. On the other hand, its overexpression leads to a
decreased TG concentration. ApoAV is synthesized in the
liver.57 In the Australian Genetic Epidemiology of
Metabolic Syndrome (GEMS) family network study, the
most common haplotype was associated with low TG and
high HDL-C.58 Two other haplotypes were associated
with an opposite prole.
The recent large-scale Mendelian randomization exper-
iment has conrmed the association of 21131T . C
APOA5 gene promoter with lipid values and CHD risk.59
Each C allele was associated with a 16% increase in TG com-
pared with common homozygotes.
Apolipoproteins C
ApoCIII is an 8800-Da glycopeptide synthesized mostly in
the liver and also in the intestine. There are three isoforms
of apoCIII characterized by a different degree of sialylation:
0, 1 and 2. ApoCIII-1 and -2 comprise 90% of apoCIII in the
plasma. APOC3 gene is part of the APOA1/C3/A4/A5 gene
cluster.60,61
Apolipoproteins C participate in the assembly of VLDL
and chylomicrons.
ApoCI is present in TG-rich lipoproteins and displaces
apoE in lipid emulsions. ApoCII, on the other hand, acti-
vates LPL and therefore contributes to VLDL lipolysis.
ApoCIII cycles between lipoproteins: chylomicrons
acquire apoC and apoE from other lipoproteins. ApoCI
and apoCIII inhibit LPL, and probably also HTGL. They
may also inhibit the uptake of remnant particles by inhibit-
ing apoE binding to receptors. ApoAV may counteract the
effects of apoCIII.
Apolipoprotein E
ApoE is a single-chain glycoprotein of 299 amino acids with
a molecular weight of 34,200 Da.62,63 It is widely distributed
across lipoprotein classes: it is present in chylomicron rem-
nants, mature VLDL, VLDL remnants, LDL and HDL. It
binds with a high afnity to the LDL-receptor as well as
to the LRP (the binding is mediated by apoCI). It facilitates
clearance of chylomicron and VLDL remnants.
Approximately 60% of plasma apoE is present in the
HDL, which exchange it with other lipoproteins. It is also
synthesized by macrophages (including macrophages
present in atherosclerotic lesions)63 and by brain astro-
cytes.64,65 ApoE controls cholesterol efux from cells
together with apoAI.66 It also has antioxidant properties
and plays a role in the regulation of inammatory response.
ApoE exists in three isoforms, E2, E3 and E4, coded by
alleles e 2, e 3 and e 4.67 The differences between the isoforms
are due to amino acids at positions 112 and 158. ApoE3 (fre-
quency 60 70% in the general population) has cysteine at
position 112 and arginine at position 158, apoE4 (frequency
505
15 20%) has arginine 112 and arginine 158, and apoE2 (fre-
quency 5 10%) has cysteine at both 112 and 158.
Individuals with e 4 allele have higher plasma cholesterol
concentration than those with e 2.68 Such individuals are
also characterized by more efcient cholesterol absorption
and increased apoB synthesis. Their apoB100 metabolism
is slower. ApoE activates LPL, HTGL and LCAT. ApoE3
and apoE4 have similar afnity to the LDL receptor, while
apoE2 has lower afnity. ApoE-decient mice show
increased plasma cholesterol concentration, and over-
expression of apoE in transgenic rabbits and mice decreases
plasma cholesterol. The e 2/2 genotype is associated with
familial dysbetalipoproteinaemia (also known as type III
hyperlipidaemia). However, although one per cent of the
population possess this genotype, dysbetalipoproteinaemia
is present in only one person in 10,000.69 On the other
hand, more than 90% of patients with dysbetalipoproteinae-
mia have e 2/2 genotype, and genotyping is used as a con-
rmation of diagnosis (see below).
Plasma apoE explains 20 40% of the variability of TG
concentrations in humans. Overexpression of apoE leads
to hypertriglyceridaemia through stimulation of VLDL pro-
duction and decrease in its clearance (for a review see
Huang).70
Apolipoprotein (a) and lipoprotein (a)
Lp(a) consists of apolipoprotein (a) (apo(a)) attached to
apoB100 by a disulphide linkage.71 An important character-
istics of the apo(a) is its structural polymorphism.72 Apo(a)
contains multiple kringles (kinks in the polypeptide chain),
and the so-called kringle 4 is the most polymorphic one of
these. Apo(a) is synthesized in the liver and it may undergo
post-translational modications. Its molecular mass ranges
between 187,000 and 800,000 Da and its plasma concentration
ranges from 0.1 to 1000 mg/dL. Its synthesis is controlled by a
series of autosomal alleles. The rate of synthesis is the main
determinant of its concentration in plasma.
Lp(a) binds to the LDL receptor. It is removed by the liver
in rodents but in humans it is also partly cleared by the
kidney. Thus, Lp(a) concentration is elevated in renal
failure. CVD risk associated with Lp(a) is also associated
with high concentrations of LDL-C. Lp(a), similarly to the
LDL, can penetrate vascular walls and associate with
intimal proteoglycans.73 Lp(a) is structurally related to plas-
minogen and potentially can interfere with its action.74,75
Although its protease activity appears to be minimal, it
might inuence the activation of plasminogen and thus
affect brinolysis. Lp(a) has been shown to inhibit tissue
plasminogen activator.76 Its concentration in plasma corre-
lates with concentrations of brinogen and D-dimer.
Clinical applications of apolipoprotein
measurements
Apolipoproteins A and B and cardiovascular risk
The plasma concentration of apoB is positively associated,
and that of apoAI inversely associated, with cardiovascular
risk.77 The apoB/apoAI ratio has been interpreted, similarly
506 Annals of Clinical Biochemistry Volume 48 November 2011
................................................................................................................................................
to the TC/HDL-C ratio, as a reection of the balance between
the atherogenic potential of the VLDL, IDL and LDL, and the
antiatherogenic effect of HDL. Current debate focuses on
whether the measurements of apoB and apoAI should comp-
lement (or substitute) measurements of TC, LDL-C and
HDL-C as markers of cardiovascular risk. There is ample,
although not completely uniform, evidence that apoB
concentration is a better marker of cardiovascular risk
than TC or LDL-C.78 A major piece of evidence was provided
by the Apolipoprotein-Related Mortality Risk Study
(AMORIS),79,80 which included 175,553 individuals followed
up for approximately 65 months. The relative risk (RR) of
fatal myocardial infarction (MI) associated with 1 standard
deviation (SD) increase in apoB concentration was about
2.7. This increased to 3.6 in individuals younger than 70
years. The apoB/AI ratio was associated with an even
higher RR of almost 4. ApoB/AI ratio, but not TC or TG,
retained predictive power after the age of 70 years.
Particularly interesting was the observation of a high predic-
tive power of the ratio in women older than 70 years, where
RR reached values between 8.0 and 9.1.
Another major study, INTERHEART, was based on 12,461
cases and 14,637 controls from 52 countries. It showed that
apoB/A1 ratio was related to the incidence of MI81 with RR
1.59 (95% CI 1.53 1.64) per 1 SD increase. Results were con-
sistent across age, gender and ethnicity. In this study, TC lost
its predictive power in patients aged 70 and older, whereas
apoAI, apoB and apoB/A1 remained predictive. The RR
was also consistently higher for the apoB/AI than for TC/
HDL-C ratio. The RR for MI associated with 1 SD change
in each of the parameters was 1.16 for TC, 0.85 for HDL-C,
1.21 for non-HDL-C, 0.67 for apoAI, 1.32 for apoB, 1.17 for
TC/HDL-C and 1.59 for apoB/apoAI. Importantly, a high
apoB/AI was associated with the highest risk, even if TC/
HDL-C ratio was relatively low.
The results of the Copenhagen City Heart Study also
suggest that apoB predicts ischaemic cardiovascular
disease better than LDL-C.82 The study involved 9231
asymptomatic women and men followed prospectively for
eight years. For apoB, the upper versus the lower tertile
hazard ratios (HR) for CHD were 1.8 (1.2 2.5), for MI 2.6
(1.4 4.7) and for any ischaemic cardiovascular event 1.8
(1.3 2.3). ApoB showed higher predictive ability than
LDL-C for CHD, MI or any ischaemic event as well as
any non-fatal ischaemic event.
In the earlier Quebec Cardiovascular Study,83 after ve
years follow-up, the RR of CHD associated with a 1 SD
increase in apoB concentration was 1.4. The RR associated
with apoAI was 0.85 but adjustment for selected lipids
and lipoproteins eliminated this association. In that study,
the apolipoproteins were measured using the now largely
abandoned Rocket immunoelectrophoresis method.
In the Third National Health and Nutrition Examination
Survey Mortality Study (NHANES III) involving 7594
American men and women (mean age 45, followed up on
average by 124 person months), the apoB measurements
predicted death (HR 1.98, CI 1.09 3.61).84 ApoAI was nega-
tively associated with risk (HR 0.48 [CI 0.27 0.85]). TC was
associated with mortality to a lesser extent (HR 1.17 [CI
1.02 1.34]). HDL-C showed a HR of 0.68 (CI 0.45 1.05),
which was borderline-signicant. ApoB/AI ratio showed a
HR of 2.14 (CI 1.11 4.10) and TC/HDL-C ratio demon-
strated a HR 1.10 (CI 1.04 1.16). After adjustment for
other risk factors, only apoB with HR 2.01 (CI 1.05 3.86)
and apoB/AI ratio with HR 2.09 (CI 1.04 4.19) remained
signicant predictors of CHD death.
On the other hand, in the MONICA/KORA Augsburg
study of men and women followed up for 13 y, the predic-
tive power of apoB/AI and TC/HDL-C was similar.85
In 2009, the Emerging Risk Factor Collaboration (ERFC),
a group of investigators with access to data from 122
previously conducted prospective studies, performed a
meta-analysis comparing the value of NHDL-C and HDL-C
testing with measurements of apoAI and apoB in the assess-
ment of vascular risk.86 They analysed population-based
studies of persons with no history of cardiovascular events.
However, they excluded AMORIS because of the lack of base-
line data on several risk factors. This meta-analysis combined
studies that measured apolipoproteins using different method-
ologies (Table 2). The mean age of participants was 58 years,
40% were women and 60% men, and a median of 6.1 years
elapsed to the rst outcome. Associations with risk were
similar for NHDL-C and apoB, and for HDL-C and apoAI.
The HR was 1.50 (1.381.62) for the NHDL-C/HDL-C ratio
(taken as a statistical equivalent of TC/HDL-C) and 1.49
(1.391.60) for 1 SD increase in apoB/AI. The median
NHDL-C and HDL-C values in analysed studies would corre-
spond roughly to TC concentrations of 66.8 mmol/L (taking
into account the lowest and highest reported HDL-C concen-
trations). The ERFC authors concluded that there is epidemio-
logical equivalence of cholesterol and apolipoprotein ratios.
As far as population-wide assessment of cardiovascular risk
is concerned, they concluded that lipid assessment in vascular
disease can be simplied by measurement of either cholesterol
levels or apolipoproteins without the need to fast and without
regard to triglyceride. They suggested that judgements
regarding apolipoprotein measurements in clinical practice
should be made on the basis of factors such as cost or methodo-
logical aspects.
Apolipoprotein concentrations were also shown to predict
cardiovascular risk in patients treated with statins. In the
Air Force/Texas Coronary Prevention Study (AFCAPS/
TEXCAPS; 6605 participants with average initial TC concen-
tration but low HDL-C), apolipoproteins, but not LDL-C,
were predictive of the rst acute event.87 Moreover, only
apoB/AI and apoB predicted subsequent risk on treatment.
The American Diabetes Association and American College
of Cardiology now recommend apolipoproteins as a best
choice to assess statin treatment.88
Table 2 Methods of measurement of apolipoprotein A and B used in
clinical studies analysed by the Emerging Risk Factors Collaboration
consortium62
Method Number of studies
Immunoturbidimetry of nephelometry 16
Immunoradiometric assays 4
Immunoelectrophoresis 1
Immunochemical methods 1
Total number of analysed studies 22
 Dominiczak and Caslake. Apolipoproteins: theory and applications
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On the other hand, in the Veterans Affairs High-Density
Lipoprotein Intervention Trial (VAHIT) of brate treatment,
where the LDL-C was relatively low, neither LDL-C nor
apoB predicted CVD risk.89 Interestingly, however, in that
study the number of LDL particles was better associated
with cardiovascular risk than the LDL-C.
Apolipoproteins A, B and the metabolic syndrome
The measurements of apoAI and apoB proved useful in
assessing CVD risk in patients with metabolic syndrome
an entity primarily associated with abnormalities in the
fuel transport pathway.90 In the Attica Study of 1514 men
and 528 women,91 the apoB/A1 ratio was the best diagnostic
marker of the metabolic syndrome as dened by the
National Cholesterol Education Program Adult Treatment
Panel III (NCEP ATPIII) criteria. The RR for metabolic syn-
drome in individuals with apoB/A1 greater than 0.74 was
3.21 (CI 2.56 4.21). Similar results were obtained by Onat
et al.,92 during prospective evaluation of 1125 men and
1123 women (age 28 74 years), followed up for 5.9 years.
ApoB predicted dyslipidaemia, metabolic syndrome and,
in women, hypertension and diabetes. This was indepen-
dent of markers of central obesity and inammation.
Individuals in the top tertile of apoB concentrations had a
RR of developing the metabolic syndrome about two-fold
that of those in the bottom tertile.
Apolipoproteins A and B in the diagnosis of specific
disorders of lipoprotein metabolism
Abnormalities in apolipoprotein concentrations dene
specic disorders of lipid metabolism. Hyperapobetalipo-
proteinaemia is a condition characterized by an increased
concentration of apoB and normal TC and variable TG con-
centrations.93 De Graaf et al. 94 have developed a diagnostic
algorithm to use apoB measurements in the diagnosis of
dyslipidaemias. Sniderman and Faraj95 also reviewed data
concerning apoA and apoB and cardiovascular risk in
relation to metabolic syndrome, insulin resistance and
abdominal obesity.
Clinical utility of apoB and apoAI measurements has
been comprehensively discussed in a Special Report from
the Working Group of the Lipoproteins and Vascular
Diseases Division of the American Association for Clinical
Chemistry.96 The Group suggested the use of apoB
measurement along with LDL-C to assess the LDL-related
risk. American Diabetic Association and American College
of Cardiology consensus also suggests that apolipoprotein
measurements are important in the metabolic syndrome.97
In a recently published roundtable discussion, there was
agreement that apoB assessments should be used by clinical
lipidologists and be incorporated into guidelines.98
Apolipoprotein B48 as a marker of postprandial
metabolism and its relation to cardiovascular risk
Humans spend a substantial proportion of their time in the
postprandial state. Zilversmit99 in 1979 suggested involve-
ment of postprandial metabolism in atherogenesis. Later,
507
Patsch100 demonstrated that individuals with CHD present
with more prolonged postprandial lipaemia. Postprandial
lipaemia can predict atherosclerosis.101 Non-fasting TG
were also a signicant predictor of CVD.102,103 Plasma con-
centration of apoB48 correlated with postprandial lipae-
mia.104 Importantly, postprandial elevation of TG,
chylomicrons and chylomicron remnants, and also of
apoB48, occurs in obesity, insulin resistance and diabetes
mellitus. This further emphasizes clinical relevance of the
fuel transport pathway and its potential markers.
Apolipoprotein CIII and cardiovascular risk
ApoCIII was found to predict cardiovascular risk. In the
Turkish population, total apoCIII, as well as non-
HDL-apoCIII and HDL-apoCIII, predicted the metabolic
syndrome with a RR of 2.5. In that study, the total apoCIII
and non-HDL-apoCIII were also predictive of the incident
CHD.105 Evidence linking apoCIII measurement with cardi-
ovascular risk has recently been reviewed by Chan et al. 61
Apolipoprotein E and cardiovascular risk
Bennet et al. 68 reviewed studies on apoE and CVD risk con-
ducted between years 1970 and 2007, analysing data from
86,067 healthy participants and using the e 3/e 3 genotype
as a reference point. Individuals with e 2/e 3 genotype
compared with e 3/e 4 had cholesterol lower by 6 9%.
Individuals with e 2/e 2, compared with e 4/e 4, had approxi-
mately 14% lower cholesterol concentration. The differences
in HDL-C between genotypes (both between e 2/e 3 and e 3/
e 4, and between e 2/e 2 and e 4/e 4) were approximately 5%.
There was a non-linear association between the e 2 genotype
and plasma TG, with the highest levels in e 2/e 2 and lowest
in e 3/e 3. When the e 3/e 3 genotype was used as a reference,
e 2/e 2 showed odds ratio (OR) for coronary risk of 0.83, e 2/
e 3 0.82, e 2/e 4 0.93, e 3/e 4 1.05 and e 4/e 4 1.22.
Apolipoprotein E and obesity
The Atherosclerosis Risk in Communities (ARIC) study
demonstrated that the body mass index progressively
increased in persons with isoforms E4 to E3 to E2.106 Also,
in obese men, apoE4 isoform was associated with higher
plasma insulin and glucose concentrations.107,108 On the
other hand, apoE-decient mice had reduced body weight
and impaired adipose tissue TG and lipid clearance.85,109,110
In other strains of mice, apoE expression worsened glucose
tolerance and decreased insulin sensitivity and facilitated
obesity.111 Inactivation of the LRP resulted in a reduction
in body weight parallel to the decreased postprandial lipid
clearance.112 ApoE, because of its role in the metabolism
of chylomicron remnants, may be particularly important
in diet-induced weight gain. Little is known about the
mechanisms linking apoE expression and weight gain in
humans. It may participate in the differentiation of preadi-
pocytes. Animal studies suggest that the most important
candidate mechanism is the facilitation of delivery of
dietary lipids to the adipose tissue, a process that may be
impaired in either the absence of apoE expression or
508 Annals of Clinical Biochemistry Volume 48 November 2011
................................................................................................................................................
defects in relevant receptors. Thus apoE, which is recog-
nized to be atheroprotective, might be a molecule that facili-
tates obesity (for a review see Kypreos et al.).62
Apolipoprotein E in the brain
ApoE concentration in the brain is secondary only to that in
the liver. It is synthesized by astrocytes and microglia, and
to a lesser extent by neurones. Its cellular uptake in the
central nervous system (CNS) is mediated by the LDL recep-
tor and by LRP. It affects growth and repair of CNS cells, is
involved in anti-inammatory response and is an antioxi-
dant. Individuals of e 4/e 4 genotype are at a greater risk
of developing a sporadic form of Alzheimers disease
(AD).113 The e 4 isoform is also a risk factor for mortality
from intracerebral haemorrhage.114
Lipoprotein (a) and cardiovascular risk
Lp(a) concentration is almost entirely genetically deter-
mined. Lifestyle factors have virtually no inuence, with
the exception of alcohol intake (heavy alcohol intake
lowers Lp(a) concentration). Catena et al. 115 reviewed
Lp(a) and cardiovascular risk in the context of hypertension.
Interestingly, Lp(a) was the best discriminator of the pres-
ence of hypertensive organ damage.116 This was related to
low molecular weight apo(a) isoforms.115
Meta-analysis by Danesh et al. 117 and by the ERFC118 also
conrmed the association between Lp(a) and CVD risk.
There was a continuous, independent, modest association
of Lp(a) with risk of CVD and stroke. On the other hand,
studies where such association was not observed include
the Physicians Health Study, the Helsinki Heart Study
and the Quebec Cardiovascular study. The discrepancies
have been ascribed to collection and sample storage pro-
blems as well as to the lack of assay standardization.
The recent Reykjavik study included 8888 male and 9681
female participants with no previous history of MI.119 The
nested controls were matched by recruitment year, sex
and age. Lp(a) was measured using a monoclonal
anti-Lp(a) antibody for capture and polyclonal anti-apoB
antibody for detection, and was not sensitive to Lp(a) iso-
forms. An increase in Lp(a) was associated with the risk
of CHD (OR 1.6 1.7; comparison between the bottom and
top tertiles of distribution): this was a lower odds ratio
than that observed for TC but higher than that for high-
sensitivity C-reactive protein or TG concentrations.
Importantly, the RR increased with increasing Lp(a) concen-
trations. The authors also performed a valuable
meta-analysis of 31 previous studies, with 14 of them per-
formed after completion of the earlier meta-analysis by
Danesh et al. The reported mean concentrations of Lp(a)
varied from 1 to 30 mg/dL. There was a consistent associ-
ation of Lp(a) with CHD risk (OR between 1.25 and 2.5
between bottom and top third of Lp(a) concentrations). In
ve studies it was between 1 and 1.25, and in three
studies there was no association.
Another systematic review of 40 studies involving 58,000
participants concluded that people with smaller apo(a) iso-
forms have an approximately two-fold higher risk of CHD
or ischaemic stroke than those with larger ones.120 Further
studies are needed to determine whether the impact of
smaller apo(a) isoforms is independent from Lp(a) concen-
tration and other risk factors.
During the European Atherosclerosis Society (EAS)
Congress in 2010, the EAS Consensus Panel reported key
points of the forthcoming scientic consensus statement
on Lp(a). They recommend that patients with moderate or
high CVD risk should be screened for Lp(a) and that there
should be a treatment goal of Lp(a) concentration below
50 mg/dL.121 This approach has been recently criticized.122
In view of the consistency of data linking Lp(a) with CVD
risk, and the long-term stability of Lp(a) concentrations, we
recommend that Lp(a) measurement should be part of the
initial assessment of the CVD risk in clinical practice.
Table 3 summarizes changes in the concentrations of plasma
apolipoproteins seen in different types of dyslipidaemia.
Methodology of apolipoprotein
measurements
ApoAI and apoB measurements are carried out by immuno-
logical techniques using antibodies raised against specic
epitopes. In early years, techniques included radial immu-
nodiffusion, radioimmunoassay and electroimmunoassay.
With the emergence of automation and advances in the pro-
duction of monospecic and monoclonal antibodies, tech-
niques used today include enzyme-linked immunosorbent
assays (ELISA), immunonephelometry and imunoturbidi-
metry. About 20 years ago, the International Federation of
Chemistry and Laboratory Medicine (IFCC) initiated a pro-
gramme to develop suitable reference material for apoAI
and B. This resulted in World Health Organization (WHO)
International Reference Materials for apoAI and
apoB.123,124 Today all instruments and assay kits should
have calibrators that can be traced to the WHO-IFCC
Reference Material. The between-assay precision on
human samples in the IFCC standardization project was
4.0 6.7% for apoAI and 4.8 8.7% for apoB.123
Individual commercial reagent kits may show interference
with lipaemia and hyperbilirubinaemia: these interferences
are noted in the technical data sheets. It is general practice
for upper limits to be quoted, e.g. it is common to see interfer-
ence at TG . 800 mg/dL (approx 9 mmol/L). Although the
precision of currently available apolipoprotein measurements
is usually below 5%, values still vary between laboratories,
and external quality assurance systems show coefcient of
variation of up to 10%. This is due to bias in different method-
ologies such as nephelometry or immunoturbidimetry, and
the use of different clinical chemistry analysers. With advances
in standardization, precision and automation, it is viable for all
clinical biochemistry laboratories to measure apoAI and apoB
routinely. We, however, recommend that each laboratory
establishes individual reference ranges specic for the instru-
ment, assay kits and population served by the laboratory.
This may be difcult to do in practice for individual
laboratories, but is possible with the help of specialist lipid lab-
oratories. In the UK, there is a network of six laboratories in the
Supra-Regional Assay Service for Cardiovascular Biomarkers
 Dominiczak and Caslake. Apolipoproteins: theory and applications 509
................................................................................................................................................

Table 3 Apolipoproteins in hypercholesterolaemia, hypertriglyceridaemia and in mixed dyslipidaemia

Atherogenic
Mixed lipoprotein
Apo Hypercholesterolaemia dyslipidaemia phenotype Hypertriglyceridaemia Usefulness as marker
AI Variable Variable Low Often low Inverse correlation with
CVD risk
CIII Correlation with CVD risk
when measured in
lipoprotein fractions
CII Rare apoCII deficiency No
causes severe
hypertriglyceridaemia
CI No
B100 High in FH, FDB, polygenic High in FCHL High marker of Usually normal Strong correlation with
hypercholesterolaemia. small dense CVD risk
Note: in LDL
hyperapobetalipoproteinaemia,
apoB100 is high
in the presence of normal TC
B48 High in LPL deficiency Marker of postprandial/
chylomicron
metabolism.
Potential marker of
CVD risk
E Genotype/isoforms related to plasma Genotyping useful in Yes genotyping/
cholesterol concentration diagnosis of phenotyping of
familial isoforms.
dyslipidaemia Atheroprotective
Lp(a) Variable Variable Variable Variable Positive correlation with
CVD risk

FH, familial hypercholesterolaemia; FDB, familial defective apoB; TC, total cholesterol; FCHL, familial combined hyperlipidaemia; CVD, cardiovascular disease
Please see text for references

(http.//www.sas-centre.org). They offer advice on the analysis
and clinical interpretation of a wide range of specialist diag-
nostic tests for lipids and lipoproteins.
Similarly to apoAI and apoB, apoAII, apoCII, apoCIII and
apoE (see below) were historically measured by radial
immunodiffusion, radioimmunoassay and electroimmu-
noassay.125 Today, measurements of these apolipoproteins
are mostly used for research purposes. Many research lab-
oratories use sodium dodecyl sulphate-polyacrylamide gel
electrophoresis (SDS-PAGE). However, for quantitation,
ELISA, immunonephelometry and imunoturbidimetry are
the methods of choice. Turbidimetric assay of apoCIII is
affected by storage (freezing). There are currently no refer-
ence methods or standards available; therefore, it is difcult
to compare values between different populations and
studies. Again, it is important that every laboratory estab-
lishes its own reference ranges, as those quoted in the tech-
nical data sheet may refer to a different population.
Methods of assessment of postprandial metabolism
The difculty in the assessment of postprandial metabolism
has been a relatively complex methodology. Measurement
of chylomicrons has been notoriously difcult because of
their short lifespan and necessity to differentiate them
from the VLDL particles. Differentiating between chylomi-
cron remnants and VLDL remnants is also difcult. Two
methodologies can be considered here: measurements of
apoB48 and of the remnant-like particles (RLP).
Conventional methods of chylomicron assessment
involve ultracentrifugation: they are separated as particles
with density below 1.006 kg/L. This can be combined
with separation of apoB48 by PAGE. Chylomicrons can
also be measured (without ultracentrifugation) in whole
plasma by SDS-PAGE, followed by measurement of
apoB48 using immunoblotting with anti-apoB48 antiserum
and ELISA assays.126 There are, however, problems with
sample stability and preparation, and the method is techni-
cally difcult and time-consuming.127 In the last few years,
highly sensitive ELISA assays became commercially avail-
able for accurate measures of apoB48.128 However, these
assays have not been standardized.
Methods of measurement of the remnant particles
Remnant lipoproteins include chylomicron and VLDL rem-
nants.129 It is generally accepted that remnant lipoproteins
are an apoE-rich fraction within TG-rich lipoproteins.
Their short half-life has hampered the development of
detection methods.
Currently, two commercially available assays are avail-
able. Both isolate lipoprotein fractions that are cholesteryl
ester- and apoE-rich, and are sensitive to freeze-thawing
and long-term storage. The assay for RLP-cholesterol
(RLP-C) has been in use in the USA since 1993. In this
assay, two monospecic monoclonal antibodies to
apoB100 and apoAI are coupled to Sepharose 4B. These
antibodies do not recognize apoE. After the non-remnant
fraction is bound by the immunoafnity gel, the fractions
are separated either by low speed ultracentrifugation or by
shaking, and RLP-C (or TG) can be measured in the
unbound fraction.130 132 Cholesterol can be measured by
510 Annals of Clinical Biochemistry Volume 48 November 2011
................................................................................................................................................
conventional means in a clinical chemistry analyser. Using
this assay, RLP-C has been reported to be an independent
risk factor in the Framingham Study.133 RLP were also elev-
ated in obesity,134 insulin resistance,135 and in patients with
diabetes and CVD.136
However, it has not been adopted widely into routine
clinical biochemistry laboratories due to the manipulations
required for the afnity gel pretreatment.
A homogeneous assay for remnant lipoprotein cholesterol
(RemL-C) has been developed. The method uses a detergent
( polyoxyethylene-polyoxybutylene block co-polymer, POE-
POB) which specically modies VLDL remnants and IDL,
and phospholipase D which modies chylomicron rem-
nants. This treatment makes cholesterol contained in these
lipoproteins (but not in other lipoprotein particles) available
to cholesterol esterase and cholesterol oxidase.
The assay allowed the measurements of cholesterol in
chylomicron remnants, VLDL remnants and IDL with no
sample pretreatment.137 Within the last few years, an auto-
mated RemL-C assay has become available, which may be
used in routine clinical chemical analysers.138
The two assays have been compared and show signicant
correlations, although RemL-C values tend to be higher than
RLP-C. The RemL-C assay was found to be more closely
associated with VLDL2 and IDL than the immunoseparation
RLP assay.138,139 It has been suggested that the discrepancies
arise because RLP values poorly reect the smaller size rem-
nants (IDL). Signicantly higher correlations have been
observed between plasma TG and RLP postprandially,
when compared with the fasting state.140 This has not been
shown yet for RemL-C. Schaeffer141 recently commented on
limitations of the RemL-C assay. He reported that RemL-C
was not a predictor of CHD risk in the Framingham
Offspring Study cohort, either prospectively or on a case-
control basis. Furthermore, there was a strong correlation
with TG (r 0.93, P , 0.001). The correlation between
RLP-C and TG was lower (r 0.79, P , 0.001) and RLP-C
was found to be an independent risk factor.133 On the other
hand, the Rem-L assay may be useful in identifying patients
with increased IDL concentrations. More studies need to be
carried out to establish the association of RemL-C with
CHD risk.
The methods for assessment of postprandial metabolism
were reviewed by Su et al. 46 They also include the measure-
ments of retinyl esters, kinetic studies using chylomicron-
like emulsions and 13C breath tests. The authors suggested
that postprandial lipaemia might be best assessed using a
combination of either serial TG measurements in response
to a standard fat load or day-long capillary TG measure-
ments, with apoB48 quantitation by the ELISA method.
Apolipoprotein E isoforms: phenotyping and
genotyping
Phenotyping
Initially, detection of apoE polymorphisms was performed
by measuring relevant proteins.142 The three isoforms
were separated by isoelectric focusing, followed by transfer
to nitrocellulose by Western blotting. The individual apoE
bands were visualized using monoclonal or polyclonal anti-
bodies to apoE,143 or direct immunoxation.144
The separation patterns are attributable to the variations
in charge. Usually migration toward the anode is faster
for the E2 isoform (Cys112/Cys158) than the E3 (Cys112/
Arg158) and E4 isoforms (Arg112/Arg158) because arginine
has a positive charge and cysteine is uncharged. However,
factors such as post-translational modications can affect
the results of apoE phenotyping. A typical Western blot
showing common ApoE isoforms is shown in Figure 5.
Genotyping
Today, as most laboratories have access to molecular tech-
niques and automated equipment, apoE isoforms are
detected by genotyping. DNA is subjected to polymerase
chain reaction using apoE primers. After digestion by
nucleases (HhaI endonuclease),145 fragments are separated
on polyacrylamide gels. Genotyping can be performed on
whole blood. Capillary gel electrophoresis can also be
used.146
There are ethical issues associated with detection of the
e 4/e 4 genotype due to its association with AD. Arguably,
the result of genotyping by a lipid laboratory should be
reported as consistent (or not consistent) with familial
dyslipidaemia.2,147
Phenotype genotype non-concordance has been repeat-
edly reported. This could be a result of post-translational
modication of the protein, such as glycation in subjects
with diabetes, or could be due to mutations in the apoE
gene resulting in protein variants, which affect the charge
of amino acids, and thus migration on the isofocusing gel.
However, not all mutations would lead to functional differ-
ences in apoE. It is important to remember that the reaction
of apoE with the LDL receptor family is dependent on a
positive charge at apoE residues 130 150, and that inter-
action with arterial wall proteoglycans depends on the
alpha-helical character of the carboxy-terminal of apoE,
which contains the heparin-binding domain. We suggest
that apoE phenotyping still has a role to play in the identi-
cation of individuals at risk for dyslipidaemia when
mutations in the apoE gene are identied.148
Measurement of apolipoprotein E in plasma
To date, research into apoE has concentrated on the investi-
gation of isoforms rather than plasma concentration of apoE.
Variation in the plasma concentration of apoE is to a large
extent genetically determined.70 ApoE2 individuals have
higher plasma levels due to slower clearance of lipoprotein par-
ticles, and apoE concentrations in E4 individuals are lower as a
result of more efcient clearance. However, within each geno-
type there is individual variation in apoE concentration.
Plasma apoE concentrations are also affected by diet. Plasma
apoE can be measured by sandwich ELISA assay,149 and by
more labour-intensive electroimmunoassay.150
Schiele et al. 150 developed extensive reference ranges for
plasma apoE, studying the French Stanislas Cohort. In this
study, apoE was measured by electroimmunoassay. Total
 Dominiczak and Caslake. Apolipoproteins: theory and applications
................................................................................................................................................
ApoE
2/2 4/2
phenotype
Figure 5 Typical Western blot showing common apoE isoforms. The order of migration towards the anode is E2 . E3 . E4
plasma apoE concentration, as well as its concentrations in
non-apoB and apoB-containing fractions, were affected by
age, gender, the common apoE polymorphisms, puberty,
serum lipid concentrations and alcohol consumption. It
was suggested that the concentration of apoE in apoB-
containing lipoproteins (remnants) is more informative
than the total plasma apoE.
Interesting data on plasma concentration of apoE came
from research on AD. Van Vliet et al. 151 assessed the risk of
late-onset AD in a study of middle aged children of
persons with AD (mean age 49.8 and 51.6 years in the AD
group and control group, respectively). They found, not sur-
prisingly, that individuals with parental history of AD were
more likely to be 14 allele carriers. Interestingly, mean
plasma apoE concentration decreased from e 2 to e 3e 3 to e 4
carriers. Individuals with parental history of AD had lower
plasma apoE concentrations than individuals without such
history.
There is no clear evidence at present to suggest that apoE
concentration should be measured routinely, although it
may have a place in the elderly. Whereas the association
between the apoE genotype and CVD is well documented,
the risk associated with circulating apoE concentration is
not known. In a study of 85-year-old individuals, it was
observed that those with high levels of apoE were at an
increased risk of cardiovascular death, independent of
their apoE genotype.149
The reason for limited use of plasma measurements might
be the distribution of apoE across lipoprotein subfractions,
including HDL. Consequently, it is relatively difcult to
explain changes in its total plasma concentration, although
the decreased clearance of TG-rich lipoproteins remains a
dominant cause of increased concentration.
Measurement of lipoprotein (a)
The size variation of the number of apo(a) kringles chal-
lenges the measurement of Lp(a) by immunochemical
methods. Problems arise in the choice of apo(a) size as the
assay calibrator and the reactivity of antibodies directed to
different kringle sizes. As a result, assays either underesti-
mate or overestimate the quantity of circulating Lp(a).
The IFCC led the development of an international refer-
ence material intended for the transfer of an Lp(a) concen-
tration to manufacturers master calibrators (IFCC
511
4/3 3/3 4/3 3/2 (-)
4
3
2
(+)
Standard Reference Material, SRM2B). The assigned value
is traceable to the consensus reference method for Lp(a).
The proposed reference material was tested in the year
2000. When used as a calibrator, no uniformity of results
was achieved for isoform-sensitive methods.152 Currently,
the only commercially available assay that is not affected
by kringle IV type-2 repeats is the one used to measure
Lp(a) in the Womens Health Study.74,153 The authors rec-
ommend that this be the assay of choice.
Concentration of apolipoproteins in different
lipoprotein subfractions
Apolipoprotein concentration in specic lipoprotein fractions
might be a good marker of CVD risk, although such measure-
ments are not suitable for routine use. Combining lipoprotein
separation and apolipoprotein measurements yielded particu-
larly interesting results in the Cholesterol and Recurrent
Events (CARE) trial.154 CARE was a study of pravastatin treat-
ment in 4159 patients with a history of acute MI and a ve-
year follow-up. VLDL-apoB, VLDL-C and VLDL-TG were
identied as predictors of recurrent coronary events.
VLDL-apoB (but not VLDL-C) was a strong predictor (with
RR 3.2 between the lowest and highest quintile). The RR for
LDL-C was 1.7 and HDL-C and plasma TG were not signi-
cant predictors. ApoCII in VLDL-plus-LDL was a strong pre-
dictor with RR 2.25, as was the apoE in HDL, with univariate
RR 2.05. Risk associated with TG was explained
by VLDL-apoB and the sum of apoCIII concentrations in
VLDL and LDL. In other studies, the sum of apoCIII
in VLDL and LDL was also increased in survivors of MI.155
ApoE in the plasma or its concentration in
VLDL-plus-LDL was associated with atherosclerosis
maybe only as a marker of apoCIII. HDL-apoE identied
patients who underwent coronary artery bypass grafting.156
The measurements of apolipoproteins in lipid subfrac-
tions emphasize one important point: the relative crudeness
of routinely used tests such as TC and in particular TG in
relation to CVD risk. Measurement of apolipoproteins in
lipid subfractions appears to be a more specic marker of
risk. However, there is a major methodological barrier
the extra step of lipoprotein separation. It is therefore unli-
kely that such methods will be used outside research
laboratories.
512 Annals of Clinical Biochemistry Volume 48 November 2011
................................................................................................................................................
The future
In the era of the omics, rapid advances are being made in
the detection and quantitation of biomarkers. Mass spec-
trometry has the potential to detect multiple proteins in a
small volume of sample at a high throughput rate. Such
an application for apolipoproteins has been reported using
ultra-performance liquid chromatography/tandem mass
spectrometry.157 This showed results for apoAI that were
comparable with those obtained in a clinical analyser and
were generated in a fraction of the time. There are hurdles
to overcome such as standardization, but this technique
has the potential to revolutionize the future analysis of
apolipoproteins. Superko158 and Mora159 have recently dis-
cussed, in an interesting format, the potential applications
of a wide range of biomarkers of lipid metabolism.
Conclusions: apolipoprotein measurements
in the clinical biochemistry laboratories
The most important use of apolipoprotein measurements in
clinical biochemistry is as markers of cardiovascular risk.
The inclusion of apoB and apoAI measurements in the
assessment of risk along with LDL-C has been rec-
ommended by a number of professional bodies. We there-
fore suggest that specialist laboratories introduce apoAI
and apoB measurements, and lead the introduction of
these assays to a wider routine service. Delaying this may
create a knowledge gap between day-to-day clinical practice
and international consensus.
Measurement of Lp(a) is also an important adjunct to risk
assessment. Because of the stability of its concentration over
time, its measurement can be limited to the initial assess-
ment of risk.
Genotyping of apolipoprotein E is important in the diag-
nosis of familial dyslipidaemia.
There is increasing recognition that postprandial metab-
olism is important in atherogenesis, particularly in the
context of obesity, insulin resistance and diabetes.
This is probably the most important emerging area in the
eld of lipoprotein metabolism, and it increases the need for
better markers of the fuel transport pathway. Currently, the
measurements of apoB48 and measurements of remnant
particle cholesterol have been the most promising markers.
Finally, the assessment of cardiovascular risk is by de-
nition multifactorial. Assessments of both lipid and non-
lipid CVD risk factors carry a degree of uncertainty. This
is due to methodological issues, to difculties in precise
denition of a given risk factors severity (e.g. family
history) or to the application of binary assessments to con-
tinuous variables (diabetes, blood pressure, smoking, de-
nition of metabolic syndrome). Therefore, the compilation
of a multifactorial risk prole remains essential.
DECLARATIONS
Competing interests: MHD delivered lectures supported
by MSD, Solvay and Abbott. MJC has received research
support from Denka Seiken, Diadexus and Japanese
ImmunoResearch Laboratories. She also received
research funding from AstraZeneca, Sano-Aventis,
GlaxoSmithKline and consultancies with Sano-Aventis
and Merck.
Funding: None.
Ethical approval: Not applicable.
Guarantor: MHD.
Contributorship: MHD wrote the majority of the review
with signicant subsequent editing and analytical input
from MJC.
Acknowledgements: The authors are very grateful to Miss
Jacky Gardiner for excellent secretarial assistance.
REFERENCES
1 Baynes J, Dominiczak MH, eds. Medical Biochemistry. 3rd edn. London:
Elsevier Mosby, 2009:653
2 Durrington P. Dyslipidemia. Lancet 2003;362:717 31
3 Rifai N, Warnick GR, Dominiczak MH, eds. Handbook of Lipoprotein
Testing. 2nd edn. Washington DC: AACC Press, 2000:817
4 Dominiczak MH. Obesity, glucose intolerance and diabetes and their
links to cardiovascular disease. Implications for laboratory medicine.
Clin Chem Lab Med 2003;41:1266 78
5 Dominiczak MH. Lipids: the story so far. Br J Cardiol 1997;4:425 9
6 Dominiczak MH. Risk factors for coronary disease: the time for a
paradigm shift? Clin Chem Lab Med 2001;39:907 19
7 Olivecrona T, Bengtsson-Olivecrona G. Lipoprotein lipase and hepatic
lipase. Curr Opin Lipidol 1990;1:222 30
8 Zambon A, Hokanson JE, Brown BG, Brunzell JD. Evidence for a new
pathophysiological mechanism for coronary artery disease regression:
hepatic lipase-mediated changes in LDL density. Circulation
1999;99:1959 64
9 Rousset X, Vaisman B, Amar M, et al. Lecithin: cholesterol
acyltransferase from biochemistry to role in cardiovascular disease.
Curr Opin Endocrinol Diabetes Obes 2009;16:163 71
10 Applebaum-Bowden D. Lipases and lecithin:cholesterol acyltransferase
in the control of lipoprotein metabolism. Curr Opin Lipidol 1995;6:130 5
11 Hesler CB, Swenson TL, Tall AR. Purication and characterization of a
human plasma cholesterol ester transfer protein. J Biol Chem
1987;262:2275 82
12 Brown MS, Goldstein JL. A receptor-mediated pathway for cholesterol
homeostasis. Science 1986;232:34 47
13 Herz J. The LDL-receptor related protein: portrait of a multifunctional
receptor. Curr Opin Lipidol 1993;4:107 13
14 Rigotti A, Miettinen HE, Krieger M. The role of the high-density
lipoprotein receptor SR-BI in the lipid metabolism of endocrine and
other tissues. Endocr Rev 2003;24:357 87
15 Tabas I, Williams KJ, Boren J. Subendothelial lipoprotein retention as
the initiating process in atherosclerosis: update and therapeutic
implications. Circulation 2007;116:1832 44
16 Ross R. The pathogenesis of atherosclerosis: a perspective for the 1990s.
Nature 1993;362:801 9
17 Rader DJ, Daugherty A. Translating molecular discoveries into new
therapies for atherosclerosis. Nature 2008;451:904 13
18 Dominiczak MH. Apoproteins and lipoproteins in human plasma. In: Rifai
N, Warnick GR, Dominiczak MH, eds. Handbook of Lipoprotein Testing.
1st edn. Washington: AACC Press, 1997:1 24
19 Powell LM, Wallis SC, Pease RJ, Edwards YH, Knott TJ, Scott J. A novel
form of tissue specic RNA processing produces apolipoprotein B48 in
intestine. Cell 1987;50:831 40
20 Eisenberg S, Sehayek E. Remnant particles and their metabolism.
Baillieres Clin Endocrinol Metab 1995;9:739 53
21 Cohn JS, Marcoux C, Davignon J. Detection, quantication, and
characterization of potentially atherogenic triglyceride-rich remnant
lipoproteins. Arterioscler Thromb Vasc Biol 1999;19:2474 86
22 Mann CJ, Yen FT, Grant AM, Bihain BE. Mechanism of plasma
cholesteryl ester transfer in hypertriglyceridaemia. J Clin Invest
1991;88:2059 66
 Dominiczak and Caslake. Apolipoproteins: theory and applications
................................................................................................................................................
23 Fruchart JC, Ailhaud G. Apolipoprotein A-containing particles:
physiological role, quantication and clinical signicance. Clin Chem
1992;38:793 7
24 Lee DM, Alaupovic P, Knight-Gibson C, Bagdade JD. Apolipoprotein-B
subclasses as acceptors of cholesteryl esters transferred by CETP. Eur J
Clin Invest 2008;38:734 42
25 Havel RJ. The formation of LDL: mechanisms and regulation. J Lipid Res
1984;25:1570 6
26 Bruckert E, Hansel B. HDL-C is a powerful lipid predictor of
cardiovascular diseases. Int J Clin Pract 2007;61:1905 13
27 von Eckardstein A, Huang Y, Assman G. Physiological role and clinical
relevance of high-density lipoprotein subclasses. Curr Opin Lipidol
1994;5:404 16
28 Florentin M, Liberopoulos EN, Wierzbicki AS, Mikhailidis DP. Multiple
actions of high-density lipoprotein. Curr Opin Cardiol 2008;23:370 8
29 Austin MA, King MD, Vranizan KM, Krauss RM. Atherogenic
lipoprotein phenotype. A proposed genetic marker for coronary heart
disease risk. Circulation 1990;82:495 506
30 Austin MA, Edwards KL. Small, dense low density lipoproteins, the
insulin resistance syndrome and noninsulin-dependent diabetes. Curr
Opin Lipidol 1996;7:167 71
31 Dominiczak MH. Apoproteins and lipoproteins in human plasma. In:
Rifai N, Warnick GR, Dominiczak MH, eds. Handbook of Lipoprotein
Testing. 2nd edn. Washington DC: AACC Press, 2000:1 29
32 Friedewald WT, Levy RI, Fredrickson DS. Estimation of the
concentration of low-density lipoprotein cholesterol in plasma, without
use of the preparative ultracentrifuge. Clin Chem 1972;18:499 502
33 Liu J, Sempos CT, Donahue RP, et al. Non-high-density lipoprotein and
very-low-density lipoprotein cholesterol and their risk predictive values
in coronary heart disease. Am J Cardiol 2006;98:1363 8
34 Shai I, Rimm EB, Harkinson SE, et al. Multivariate assessment of lipid
parameters as predictors of coronary heart disease among
postmenopausal women: potential implications for clinical guidelines.
Circulation 2004;110:2824 30
35 Jiang R, Schulze MB, Li T, et al. Non-HDL cholesterol and
apolipoprotein B predict cardiovascular disease events among men
with type 2 diabetes. Diabetes Care 2004;27:1991 7
36 Blake GJ, Otvos JD, Rifai N, Ridker PM. Low-density lipoprotein
particle concentration and size are determined by nuclear magnetic
resonance spectroscopy as predictors of cardiovascular disease in
women. Circulation 2002;106:1930 7
37 Hainline AJ, Karon J, Kippel K. Manual of Laboratory Operations: Lipid
Research Clinics Program, Lipid and Lipoprotein Analysis. 2nd edn.
Washington, DC: Department of Health and Human Services, 1983
38 Marcovina S, Packard CJ. Measurement and meaning of apolipoprotein
B plasma levels. J Intern Med 2006;259:437 46
39 Durrington PN. Hyperlipidaemia: Diagnosis and Treatment. 3rd edn.
London: Hodder Arnold, 2007:66 91
40 Olofsson SO, Boren J. Apolipoprotein B: a clinically important
apolipoprotein, which assembles atherogenic lipoproteins and
promotes the development of atherosclerosis. J Intern Med
2005;258:395 410
41 Knott TJ, Pease RJ, Powell LM, et al. Complete protein sequence and
identication of structural domains of human apolipoprotein B. Nature
1986;323:734 8
42 Hussain MM, Shi J, Dreizen P. Microsomal triglyceride transfer protein
and its role in apoB-lipoprotein assembly. J Lipid Res 2003;44:22 32
43 Blackhart BD, Ludwig EM, Pierotti VR, et al. Structure of the human
apolipoprotein B gene. J Biol Chem 1986;261:15364 7
44 Innerarity TL, Mahley TL, Weisgraber KH, et al. Familial defective
apolipoprotein B-100: a mutation of apolipoprotein B that causes
hypercholesterolemia. J Lipid Res 1990;31:1337 49
45 Hardman DA, Kane JP. Isolation and characterization of apolipoprotein
B-48. Methods Enzymol 1986;128:262 72
46 Su JW, Nzekwu MMU, Cabezas MC, Redgrave T, Proctor SD. Methods
to assess impaired post-prandial metabolism and the impact for early
detection of cardiovascular disease risk. Eur J Clin Invest 2009;39:741 54
47 Berry SEE. Postprandial lipaemia the inuence of diet and its link to
coronary heart disease. Nutr Bull 2005;30:314 22
48 Bruns GA, Karathanasis SK, Breslow JL. Human apolipoprotein A-I
C-III gene complex is located on chromosome 11. Arteriosclerosis
1984;4:97 102
513
49 Feng H, Li X-A. Dysfunctional high-density lipoprotein. Curr Opin
Endocrinol Diabetes Obes 2009;16:156 62
50 Kilimov AN, Gurevich VS, Nikiforova AA, et al. Antioxidative activity
of high density lipoproteins in vivo. Atherosclerosis 1993;100:13 8
51 Cheung M, Albers JJ. Distribution of high density lipoprotein particles
with different apoprotein composition: particles with AI and AII and
particles with AI but no AII. J Lipid Res 1982;23:747 53
52 Julve J, Escola-Gil JC, Rotllan N, et al. A human apolipoprotein AII
determines plasma triglycerides by regulating lipoprotein lipase
activity and high-density lipoprotein proteome. Arterioscler Thromb Vasc
Biol 2010;30:232 8
53 Warden CH, Hedrick CC, Quiao J-H, Castellani LW, Lusis AJ.
Atherosclerosis in transgenic mice overexpressing apolipoprotein A-II.
Science 1993;261:469 72
54 Birjmohun RS, Dallinga-Thie GM, Kuivenhoven JA, et al.
Apolipoprotein A-II is inversely associated with risk of future coronary
disease. Circulation 2007;116:2029 35
55 Tailleux A, Duriez P, Fruchart JC, Clavey V. Apolipoprotein A-II, HDL
metabolism and atherosclerosis. Atherosclerosis 2002;164:1 13
56 Weinberg RB. Apolipoprotein A-IV polymorphisms and diet gene
interactions. Curr Opin Lipidol 2002;13:125 34
57 Calandra S, Oliva CP, Tarugi P, Bertolini S. APOA5 and triglyceride
metabolism, lesson from human APOA5 deciency. Curr Opin Lipidol
2006;17:122 7
58 Laurila PP, Naukkarinen J, Kristiansson K, et al. Genetic association
and interaction analysis of USF1 and APOA5 on lipid levels and
atherosclerosis. Arterioscler Thromb Vasc Biol 2010;30:346 52
59 Triglyceride Coronary Disease Genetics Consortium and Emerging
Risk Factors Collaboration. Triglyceride-mediated pathways and
coronary disease: collaborative analysis of 101 studies. Lancet
2010;375:1634 9
60 Lai C-Q, Parnell LD, Ordovas JM. The APOAI/C3/A4/A5 gene cluster,
lipid metabolism and cardiovascular disease risk. Curr Opin Lipidol
2005;16:153 66
61 Chan DC, Chen MM, Ooi EMM, Watts GF. An ABC of apolipoprotein
C-III: a clinically useful new cardiovascular risk factor? Int J Clin Pract
2008;62:799 809
62 Kypreos KE, Karagiannides I, Fotiadou EH, et al. Mechanisms of obesity
and related pathologies: role of apolipoprotein E in the development
of obesity. FEBS J 2009;276:5720 8
63 Mahley RW, Innerarity TL, Rall JSC, Weisgraber KH, Taylor JM.
Apolipoprotein E: genetic variants provide insights into its structure
and function. Curr Opin Lipidol 1990;1:87 95
64 Donahue JE, Johanson CE. Apolipoprotein E, amyloid-A, and blood
brain barrier permeability in Alzheimer disease. J Neuropathol Exp
Neurol 2008;67:261 70
65 Boyles JK, Pitas RE, Wilson E, Mahley RW, Taylor JM. Apolipoprotein E
associated with astrocytic glia of the central nervous system and with
nonmyelinating glia of the peripheral nervous system. J Clin Invest
1985;76:1501 13
66 Chroni A, Nieland TJ, Kypreos KE, Krieger M, Zannis VI. SR-BI
mediates cholesterol efux via its interactions with lipid-bound ApoE.
Structural mutations in SR-BI diminish cholesterol efux. Biochemistry
2005;44:13132 43
67 Utermann G. Apolipoprotein E polymorphisms in health and disease.
Am Heart J 1987;113:433 40
68 Bennet AM, Di Angelantonio E, Ye Z, et al. Association of
apolipoprotein E genotypes with lipid levels and coronary risk. JAMA
2007;298:1300 11
69 Brewer HB, Zech LA, Gregg RE, Schwartz D, Schaeffer EJ. Type III
hyperlipoproteinemia: diagnosis, molecular defects, pathology and
treatment. Ann Int Med 1983;98:623 40
70 Huang Y. Mechanisms linking apolipoprotein E isoforms with
cardiovascular and neurological diseases. Curr Opin Lipidol
2010;21:337 45
71 Loscalzo J. Lipoprotein(a): a unique risk factor for atherothrombotic
disease. Arteriosclerosis 1990;10:672 9
72 Utermann G, Menzel HJ, Kraft HG, Duba HC, Kemmler HG, Seitz C.
Lp(a) glycoprotein phenotypes. J Clin Invest 1987;80:458 65
73 Rath M, Niendorf A, Reblin T, Dietel M, Krebber H-J, Beisiegel U.
Detection and quantication of lipoprotein (a) in the arterial wall of 107
coronary bypass patients. Arteriosclerosis 1989;9:579 92
514 Annals of Clinical Biochemistry Volume 48 November 2011
................................................................................................................................................
74 McLean JW, Tomlinson JE, Kuang W-J, et al. cDNA sequence of human
apolipoprotein (a) is homologous to plasminogen. Nature
1987;330:132 7
75 Karadi I, Kostner GM, Gries A, Nimpf J, Romics L, Malle E. Lipoprotein
(a) and plasminogen are immunochemically related. Biochim Biophys
Acta 1988;960:91 7
76 Loscalzo J, Weinfeld M, Fless G, Scanu AM. Lipoprotein (a), brin
binding and plasminogen activation. Arteriosclerosis 1990;10:240 5
77 Andrikoula M, McDowell IFW. The contribution of apoB and apoA1
measurements to cardiovascular risk assessment. Diabetes Obes Metab
2008;10:271 8
78 Thompson A, Danesh J. Association between apolipoprotein B,
apolipoprotein A1, the apolipoprotein B/A1 ratio and coronary heart
disease: a literature-based meta-analysis of prospective studies. J Intern
Med 2006;259:481 92
79 Walldius G, Jungner I. Apolipoprotein B and apolipoprotein A1: risk
indicators of coronary heart disease and targets for lipid-modifying
therapy. J Intern Med 2004;255:188 205
80 Walldius G, Jungner I, Holme I, et al. High apolipoprotein B, low
apolipoprotein A-1, and improvement in the prediction of fatal
myocardial infarction (AMORIS study): a prospective study. Lancet
2001;358:2026 33
81 McQueen MJ, Hawken S, Wang X, et al. Lipids, lipoproteins, and
apolipoproteins as risk markers of myocardial infarction in 52 countries
(the INTERHEART study): a case-control study. Lancet 2008;372:224 33
82 Benn M, Nordestgaard BG, Jensen GB, Tybjaerg-Hansen A. Improving
prediction of ischemic cardiovascular disease in the general population
using apolipoprotein B: the Copenhagen City Heart Study. Arterioscler
Thromb Vasc Biol 2007;27:661 70
83 Lamarche B, Moorjani S, Lupien PJ, et al. Apolipoprotein A-1 and B
levels and the risk of ischemic heart disease during a 5 year follow-up of
men in the Quebec Cardiovascular Study. Circulation 1996;94:273 8
84 Sierra-Johnson J, Fisher RM, Romero-Corral A, et al. Concentration of
apolipoprotein B is comparable with the apolipoprotein B/
apolipoprotein A-I ratio and better than routine clinical lipid
measurements in predicting coronary heart disease mortality: ndings
from a multi-ethnic US population. Eur Heart J 2009;30:710 7
85 Meisinger C, Loewel H, Mraz W, Koenig W. Prognostic value of
apolipoprotein B and A-1 in the prediction of myocardial infarction in
middle-aged men and women: results from the MONICA/KORA
Augsburg Cohort Study. Eur Heart J 2005;26:271 8
86 The Emerging Risk Factor Collaboration. Major lipids, apolipoproteins,
and risk of vascular disease. JAMA 2009;302:1993 2000
87 Gotto AM, Whitney E, Stein EA, et al. Relation between baseline and
on-treatment lipid parameters and rst acute major coronary events in
the Air Force/Texas Coronary Atherosclerosis Prevention Study
(AF-CAPS/TexCAPS). Circulation 2000;101:477 84
88 Brunzell JD, Davidson M, Furberg CD, et al. Lipoprotein management
in patients in cardiometabolic risk: conference report from the
American Diabetes Association and the American College of
Cardiology Foundation. JACC 2008;51:1512 24
89 Otvos JD, Collins D, Freedman DS, et al. Low-density lipoprotein and
high-density lipoprotein particle subclasses predict coronary events and
are favourably changed by gembrozil therapy in the Veterans Affairs
High-Density Lipoprotein Intervention Trial. Circulation
2006;113:1556 63
90 Dominiczak MH. Metabolic syndrome. CPD Clin Biochem 2004;6:3 9
91 Pitsavos C, Panagiotakos DB, Skoumas J, et al. Risk stratication of
apolipoprotein B, apolipoprotein AI and apolipoprotein B/AI ratio on
the prevalence of the metabolic syndrome: the ATTICA Study.
Angiology 2008;59:335 41
92 Onat A, Hergenc G, Yazc M, Karabulut A, Albayrak S. Serum
apolipoprotein B predicts dyslipidaemia, metabolic syndrome and, in
women, hypertension and diabetes, independent of markers of central
obesity and inammation. Int J Obes 2007;37:1119 25
93 Teng B, Sniderman AD, Soutar AK, Thompson GR. Metabolic basis of
hyperapobetalipoproteinemia. Turnover of apolipoprotein B in low
density lipoprotein and its precursors and subfractions compared with
normal and familial hypercholesterolemia. J Clin Invest 1986;77:663 72
94 De Graaf J, Couture P, Sniderman A. A diagnostic algorithm for the
atherogenic apolipoprotein B dyslipoproteinemias. Nat Clin Pract
Endocrinol Metab 2008;4:608 18
95 Sniderman AD, Faraj M. Apolipoprotein B, apolipoprotein A-I, insulin
resistance and the metabolic syndrome. Curr Opin Lipidol 2007;18:633 7
96 Contois JH, McConnell JP, Sethi AA, et al. Apolipoprotein B and
cardiovascular disease risk: position statement from the AACC
Lipoproteins and Vascular Diseases Division Working Group on Best
Practices. Clin Chem 2009;55:407 19
97 Brunzell JD, Davidson M, Furberg CD, et al. Lipoprotein management
in patients in cardiometabolic risk: conference report from the
American Diabetes Association and the American College of
Cardiology Foundation. JACC 2008;51:1512 24
98 Brown WV, Myers GL, Sniderman A, Evan Stein E. Should we use apoB
for risk assessment and as a target for treatment? J Clin Lipidol
2010;4:144 51
99 Zilversmit DB. Atherogenesis: a postprandial phenomenon. Circulation
1979;60:473 85
100 Patsch JR, Miesenbock G, Hopferwieser T, et al. Relationship of
triglyceride metabolism and coronary artery disease. Arterioscler Thromb
1992;12:1336 45
101 Karpe F. Postprandial lipoprotein metabolism and atherosclerosis.
J Intern Med 1999;246:341 55
102 Bansal S, Buring JE, Rifai N, et al. Fasting compared with nonfasting
triglycerides and risk of cardiovascular events in women. JAMA
2007;298:309 16
103 Brge G, Nordestgaard BG, Benn M, et al. Nonfasting triglycerides and
risk of myocardial infarction, ischaemic heart disease, and death in men
and women. JAMA 2007;298:299 308
104 Smith D, Watts GF, Dane-Stewart C, Mamo JC. Post-prandial CM
response may be predicted by a single measurement of plasma
apolipoprotein B48 in the fasting state. Eur J Clin Invest 1999;29:204 9
105 Onat A, Hergenc G, Ayhan E, et al. Metabolism of serum apolipoprotein
C-III in high-density lipoprotein: a key diabetogenic risk factor in Turks.
Diabet Med 2009;26:981 8
106 Volcik KA, Barkley RA, Hutchinson RG, et al. Apolipoprotein E
polymorphisms predict low density lipoprotein cholesterol levels and
carotid artery wall thickness but not incident coronary heart disease in
12,491 ARIC study participants. Am J Epidemiol 2006;164:342 8
107 Elosua R, Demissie S, Cupples LA, et al. Obesity modulates the
association among APOE genotype, insulin, and glucose in men. Obes
Res 2003;11:1502 8
108 Marques-Vidal P, Bongard V, Ruidavets JB, et al. Obesity and alcohol
modulate the effect of apolipoprotein E polymorphism on lipids and
insulin. Obes Res 2003;11:1200 6
109 Huang ZH, Reardon CA, Mazzone T. Endogenous ApoE expression
modulates adipocyte triglyceride content and turnover. Diabetes
2006;55:3394 402
110 Hofmann SM, Perez-Tilve D, Greer TM, et al. Defective lipid delivery
modulates glucose tolerance and metabolic response to diet in
apolipoprotein E-decient mice. Diabetes 2008;57:5 12
111 Gao J, Katagiri H, Ishigaki Y, et al. Involvement of apolipoprotein E
in excess fat accumulation and insulin resistance. Diabetes 2007;56:
24 33
112 Hofmann SM, Zhou L, Perez-Tilve D, et al. Adipocyte LDL receptor-
related protein-1 expression modulates postprandial lipid transport and
glucose homeostasis in mice. J Clin Invest 2007;117:3271 82
113 Saunders AM, Schmader K, Breitner JCS, et al. Apolipoprotein E allele
distributions in late-onset Alzheimers disease and in other
amyloid-forming diseases. Lancet 1993;342:710 11
114 McCarron MO, Hoffmann KL, DeLong DM, Gray L, Saunders AM,
Alberts MJ. Intracerebral hemorrhage outcome: apolipoprotein E
genotype, hematoma, and edema volumes. Neurology 1999;53:2176 9
115 Catena C, Novello M, Lapenna R, et al. New risk factors for
atherosclerosis in hypertension: focus on the prothrombotic state and
lipoprotein (a). J Hypertens 2005;23:1617 31
116 Sechi LA, Kronenberg F, De Carli S, Falleti E, Zingaro L. Association of
serum lipoprotein (a) levels and apolipoprotein (a) size polymorphism
with target-organ damage in arterial hypertension. JAMA
1997;277:1689 95
117 Danesh J, Collins R, Peto R. Lipoprotein (a) and coronary heart disease:
meta-analysis of prospective studies. Circulation 2000;102:1082 5
118 The Emerging Risk Factors Collaboration. Lipoprotein (a) concentration
and the risk of coronary heart disease, stroke, and nonvascular
mortality. JAMA 2009;302:412 23
 Dominiczak and Caslake. Apolipoproteins: theory and applications
................................................................................................................................................
119 Bennet A, Di Angelantonio E, Erqou S, et al. Lipoprotein(a) levels and
risk of future coronary heart disease. Large-scale prospective data. Arch
Intern Med 2008;168:598 60
120 Erqou E, Thompson A, Di Angelantonio E, et al. Apolipoprotein(a)
isoforms and the risk of vascular disease. J Am Coll Cardiol
2010;55:2160 7
121 European Atherosclerosis Society (EAS). Consensus Panel Presentation
at the Meeting of the European Atherosclerosis Society, Hamburg, 2010
122 Goldhaber SZ. European Atherosclerosis Society screening
recommendations for lipoprotein (a) and high-sensitivity C-reactive
protein: double standard or failure of evidence-based medicine? Clin
Chem 2010;56:1544 6
123 Marcovina SM, Albers JJ, Dati F, Ledue TB, Ritchie RF. International
Federation of Clinical Chemistry standardisation project for
measurements of apolipoprotein A-I and B. Clin Chem 1991;37:1676 82
124 Marcovina SM, Albers JJ, Kennedy H, Mei JV, Henderson LO, Hannon
WH. International Federation of Clinical Chemistry standardisation
project for measurements of apolipoprotein A-I and B. IV.
Comparability of apolipoprotein B values by use of international
reference material. Clin Chem 1994;40:586 92
125 Labeur C, Rosseneu M. Immunological assays of apolipoproteins A-II,
A-IV, C-I, C-II, and C-III in plasma: methods and applications. In: Rifai
N, Warnick GR, Dominiczak MH, eds. Handbook of Lipoprotein Testing.
2nd edn.) Washington DC: AACC Press, 2000:387400
126 Smith D, Proctor SD, Mamo CLM. A highly sensitive assay for
quantitation of apolipoprotein B48 using an antibody to human
apolipoprotein B and enhanced chemiluminescence. Ann Clin Biochem
1997;34:185 9
127 Karpe F, Hamsten A. Determination of apolipoproteins B-48 and B-100
in triglyceride-rich lipoproteins by analytical SDS-PAGE. J Lipid Res
1994;35:1311 7
128 Valdivielso P, Puerta S, Rioja J, et al. Postprandial apolipoprotein B48 is
associated with asymptomatic peripheral arterial disease: a study in
patients with type 2 diabetes and controls. Clin Chim Acta
2010;411:433 7
129 Havel RJ. Remnant lipoproteins as therapeutic targets. Curr Opin Lipidol
2000;11:15 20
130 Nakajima K, Saito T, Tamura A, et al. Cholesterol in remnant-like
lipoproteins in human serum using monoclonal anti apoB-100 and anti
apoA-I immunoafnity mixed gels. Clin Chim Acta 1993;223:53 71
131 Mc Namara J, Shah PK, Nakajima K, et al. Remnant lipoprotein
cholesterol and triglyceride reference ranges from the Framingham
Study. Clin Chem 1998;44:1224 32
132 Van Hees AMJ, Saris WHM, Dallinga-Thie GMD, Hul GB, Martinez JA.
Fasting and postprandial remnant-like particle cholesterol concentration
in obese participants is associated with plasma triglycerides, insulin
resistance and body fat distribution. J Nutr 2008;138:2399 405
133 McNamara JR, Shah PK, Nakajima JM, et al. Remnant-like particle
(RLP) cholesterol is an independent cardiovascular risk factor in
women: results from the Framingham Heart Study. Atherosclerosis
2001;154:229 36
134 Chan DC, Watts GF, Barrett PH, et al. Markers of triglyceride-rich
lipoprotein remnant metabolism in visceral obesity. Clin Chem
2002;48:278 83
135 Abbasi F, McLaughlin T, Lamendola C, et al. Fasting remnant
lipoprotein cholesterol and triglyceride concentrations are elevated in
nondiabetic, insulin-resistant, female volunteers. J Clin Endocrinol Metab
1999;84:3903 6
136 Higashi K, Ito T, Nakajima K, et al. Remnant-like particles cholesterol is
higher in diabetic patients with coronary artery disease. Metabolism
2001;50:1462 5
137 Miyauchi K, Kayahara N, Ishigami M, et al. Development of
homogeneous assay to measure remnant lipoprotein cholesterol. Clin
Chem 2007;53:2128 35
138 Nagashima M, Mori Y, Morita R, et al. Remnant lipoprotein cholesterol
homogeneous assay (RemL-C) is closely associated with
very-low-density lipoprotein remnants:comparison with the
immunoseparation assay. Rinsho Byori 2008;56:973 9
139 Yoshida H, Kurosawa H, Hirowatari Y, et al. Characteristic comparison
of triglyceride-rich remnant lipoprotein measurement between a new
515
homogenous assay (RemL-C) and a conventional immunoseparation
method (RLP-C). Lipids Health Dis 2008;7:18 22
140 Nakajima K, Nakano T, Moon HD, et al. The correlation between TG vs.
remnant lipoproteins in the fasting and postprandial plasma of 23
volunteers. Clin Chim Acta 2009;404:124 7
141 Schaeffer EJ. Limitations of automated remnant lipoprotein cholesterol
assay for diagnostic use. Clin Chem 2009;55:2061 2
142 Siest G, Pillot T, Regis-Bailly A, et al. Apolipoprotein E: an important
gene and protein to follow in laboratory medicine. Clin Chem
1995;41:1068 86
143 Havekes LM, de Knijff P, Beisiegel U, Havinga J, Smit M, Klasen E. A
rapid micromethod for apolipoprotein E phenotyping directly in serum.
J Lipid Res 1987;28:455 63
144 Hackler R, Schaefer JR, Motznu S, et al. Rapid determination of
apolipoprotein E phenotypes from whole plasma by automated electric
focusing using PhastSystem and immunoxation. J Lipid Res
1994;35:153 8
145 Hixson JE, Vernier DT. Restriction isotyping of human apolipoprotein E
by gene amplication and cleavage with HhaI. J Lipid Res 1990;31:545 8
146 Siest G, Schlenck A, Starck M, et al. Apolipoprotein E: laboratory
determinations and clinical interest. In: Rifai N, Warnick GR,
Dominiczak MH, eds. Handbook of Lipoprotein Testing. 2nd edn.
Washington DC: AACC Press, 2000:40140
147 National Institute on Aging/Alzheimers Association Working Group.
Consensus statement. Apolipoprotein E genotyping in Alzheimers
disease. Lancet 1996;347:1091 5
148 Stephens JW, Sozen MM, Whittall RA, et al. Three novel mutations in
the apolipoprotein E gene in a sample of individuals with type 2
diabetes mellitus. Clin Chem 2005;51:119 24
149 Mooijaart SP, Berbea JFP, van Heemst D, et al. ApoE plasma levels and
risk of cardiovascular mortality in old age. PLoS Med 2006;3:874 88
150 Schiele F, Vincent-Viry M, Starck M, et al. Apolipoprotein E in
apolipoprotein B (apo B)- and non-apo B-containing lipoproteins in
3523 participants in the Stanislas Cohort: biological variation and
genotype-specic reference limits. Clin Chem 2002;48:291 300
151 Van Vliet P, Westendorp RGJ, Eikelenboom P, et al. Parental history of
Alzheimer disease associated with lower plasma apolipoprotein E
levels. Neurology 2009;73:681 7
152 Marcovina SM, Albers JJ, Scanu A, et al. Use of reference material
proposed by the International Federation of Clinical Chemistry and
Laboratory Medicine to evaluate analytical methods for the
determination of plasma lipoprotein (a). Clin Chem 2000;46:1956 67
153 Danik JS, Rifai N, Buring JE, Ridker PM. Lipoprotein (a), measured with
an assay independent of apolipoprotein (a) isoform size, and risk of
future cardiovascular events among initially healthy women. JAMA
2006;296:1363 70
154 Sacks FM, Alaupovic P, Moye LA, et al. VLDL, apolipoproteins B, CIII
and E, and risk of recurrent coronary events in the Cholesterol and
Recurrent Events (CARE) trial. Circulation 2000;102:1886 92
155 Luc G, Fievet C, Artweiler D, et al. Apolipoproteins CIII and E in ApoB
and non-apoB-containing lipoproteins in two populations at contrasting
risk for myocardial infarction: the ECTIM study. J Lipid Res
1996;37:508 17
156 Chivot L, Mainard F, Bigot E, et al. Logistic discriminant analysis of
lipids and apolipoproteins in a population of coronary bypass patients
and the signicance of apolipoproteins CIII and E. Atherosclerosis
1990;82:205 11
157 Kay RG, Gregory B, Grace PB, Pleasance S. The application of
ultra-performance liquid chromatography/tandem mass spectrometry
to the detection and quantitation of apolipoproteins in human serum.
Rapid Commun Mass Spectrom 2007;21:2585 93
158 Superko HR. Advanced lipoprotein testing and subfractionation are
clinically useful. Circulation 2009;119:2383 95
159 Mora S. Advanced lipoprotein testing and subfractionation are not (yet)
ready for routine clinical use. Circulation 2009;119:2396 404
(Accepted 17 June 2011)