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___ This Week’s Citation Classic ® OCTOBER 22. 1990

Albertsson P.A. Partition ofce/I particles and macromolecules.

New York: Wiley. 1196011986.
[Department of Biochemistry. University of Lund. Sweden)

This book describes the separation and purification ol wherein mixtures of hydroxylapatite and chloro-
cell particles and biomolecules by phase partition. plasts suspensions were shaken and the settling of
Aqueous mixtures ot two different polymers produce the green particles was observed. Again the chloro-
liquid-liquid two-phase systems that allow a mild and plasts were firmly adsorbed even in the presence of
selective partitioning of proteins, nucleic acids, mem- 1-2 molar (M) phosphate.
brane vesicles. cell organdies, and even whole cells. I decided to use a detergent to facilitate desorp-
[The Sd® indicates that the various editions of this tion. From a book on detergents, I remembered the
book have been cited in over 1,100 publications.l name polyethylene glycol (PEG). Since this hap-
pened to be in a bottle on the shelf, I tried it. An
—.~ .~ aqueous solution of PEG was mixed with an
hydroxylapatite sediment containing the firmly ad-
Separation of Cell Particles and Molecules sorbed chloroplast particles in about 1 M potassium
phosphate. The result was most spectacular. The
Per-Ake Albertsson intense green color of the chboroplasts, earlier so
Department of Biochemistry strongly associated with hydroxylapatite, was now
Chemical Center present in a liquid layer on top of the phosphate
4. University of Lund buffer; the hydroxylapatite turned white and was
S-221 00 Lund completely purged of chloroplasts. Owing to the
Sweden high phosphate concentration, a two-phase system
was formed and the upper, PEG-rich phase appar-
July 3, 1990 ently had a stronger affinity for the chloroplasts
than did the hydroxylapatite. Later I found that PEG
is not a detergent; it is used for thc manufacture of
The first edition of this book was my PhD thesis in certain detergents. Thus, my experiment was a re-
biochemistry at the University of Uppsala, Sweden. warding “mistake.” Due to the intense color of the
In 1954 I began my graduate studies with Arne
chioroplasts, the phenomenon was very impressive,
Tiselius. My supervisor was Hákan Leyon, an elec- and this helped me to realize that partitioning might
tron microscopist interested in the chioroplast. My
be used for the separation of cell partides.
first task was to isolate pyrenoids, characteristic
Since PEG is a polymer and somewhat hydropho-
structures of chloroplasts from green algae. Besides bic compared to potassium phosphate buffer, the
centrifugation I also used chromatography on col-
umns of hydroxylapatite, which had been intro- latter was replaced by another hydrophilic polymer,
dextran. In this way an aqueous polymer-polymer
duced by Tiselius for separation of proteins. It was
of great methodological interest to see whether two-phase system was obtained. The dextran PEG
these columns could be applied also to cell parti- phase system, which is very mild towards biological
cles. material, has since been used for many different
My experiments failed, however, due to irrevers- separation purposes.t-a It has, e.g., been success- 4
ible adsorption. In a typical chromatographic ex- fully used for purification of plasma membranes
periment with hydroxylapatite, a substance is first and for separation of inside out from right side out
adsorbed at low concentrations of phosphate buffer thylakoids or plasma membrane” vesicles. The
and then eluted at higher concentrations. The ebb- method is highly versatile and can be applied ‘uoth
roplast particles were easily adsorbed onto the col- on a small and on a large industrial scale involving
umn but could not be desorbed. The green band on several thousands of liters.’ A Japanese translation
top of the column did not budge upon elution with of my book was published in 1971; a Russian, in
various buffers. Could it be that the relatively large 1974.
particles were mechanically trapped between the However, my original problem as a PhD student
hydroxylapatite grains of the column? To eliminate in 1954, to isolate pyrenoids, still remains to be
this possibility, I switched to batch experiments solved.

t. Walter H, Brooks 0 E & Fisher 0. mIs. Partitioning in aqueous two.phase sos,ems. Orlando. FL: Academic Press,
2. Albertsson P-A. Analysis of the domain structure of membranes by fragmentation and separation in aqueous polymer
two-phase systems. Quart. Ret’. Bittphvs. 21:68-98, 1988.
3. ,Johansson G. Separation of biopolvmers by partition. Sepur. Parif. Method. 17:85-203, 988.
4. Larsson C. Widell S & Sommarin M. Inside-out plant plasma membrane vesicles of high purity obtained by aqueous
two-phase partitioning. FEBS Leo. 229:289-92. 988. (Cited tO times.)
5. Andersson B, Sundby C, Akerlund HE & Alberssson P-A. Inside-out ibylukoid vesicles. An tmportunt toot for the
characterization of the photosynthetic membrane. Physiol. Plant. 63:322.30, 1985. (Cited 20 times.)
6. Hustedt H. Kroner K H, Menge U & Kula M-R. Protein recovery using two-phase systems. Trends Btotech.
3:139-44. 1985. )Cited 25 times.)