Essential A level Biology Practical Skills

What you need to be able to do:

1. Carrying out basic laboratory techniques and understanding the principles that
underlie them
2. Working safely, responsibly and legally in the laboratory, with due attention to
ethical aspects
3. Designing, planning and conducting biological inestigations
!. "btaining, recording, collating and analysing biological data
#. $sing data in seeral forms e.g. numerical, te%tual, erbal and graphical
&. 'aluating your e%perimental technique
(eneral )pproach to *ractical Work
+ead handouts in adance, where possible, so you understand why you are doing a
particular practical and the principles behind it
,e aware of the time in which you hae to work
Consider safety ha-ards before you begin
"rganise your working bench space
Write up work as soon as possible after the practical
)ccuracy and *recision in techniques
)ccuracy . the closeness of a measured data alue to its true alue
*recision . the closeness of repeated measurements to each other
/o0 a balance with a fault in it could gie precise 1i.e. ery repeatable2 results but
inaccurate 1untrue2 results. $nless there is a bias 1fault2 in the measuring system,
precision will lead to accuracy.
,ias can be due to34
5ncorrectly calibrated instruments e.g. faulty water bath
'%perimental manipulations e.g. using a thermometer to measure temperature can
itself decrease the temperature
/ub6ectie ideas by the e%perimenter e.g. 6udging when an end4point is reached,
or fi%ing results to fit those e%pected
7inimising 'rrors
When designing an e%periment34
'nsure that the independent ariable is the only ma6or factor that changes
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Essential A level Biology Practical Skills
5ncorporate a control e%periment to show it is only the independent ariable
which causes the measured effect
Where appropriate, select e%perimental sub6ects randomly to cancel out ariation
arising from biased selection 1this is important in ecological inestigations2
8eep the number of replicates as high as possible
'nsure the same number of replicates is done for each alue of the independent
ariable
5dentify other factors which could affect the dependent ariable and keep them
constant 1control ariables2 e.g. temperature, olumes of solutions, light intensity,
time for reaction
7inimising 7easurement 'rror
Common source is carelessness e.g. reading a scale in the wrong direction9 reduce
this by more careful recording 1think how you can do this2 and by repeating the
measurement
)ccuracy depends on using suitable equipment with care.
)ccurate measurement of liquids
:igh iscosity liquids are difficult to transfer9 allow time for all the liquid to
transfer
"rganic solents or hot liquids may eaporate quickly, making measurements
inaccurate9 transfer these liquids quickly and coer containers
;iquids likely to froth e.g. yeast or protein solutions are difficult to measure9
transfer slowly
/uspensions e.g. yeast or cell cultures may sediment9 mi% well before transferring
$se measuring cylinders on a leel surface so the scale is hori-ontal9 fill to below
the desired mark, then add liquid slowly e.g. by pipette to reach desired leel
7ake sure there are no air bubbles in syringes when measuring olumes. '%pel
liquids slowly and touch the end of the syringe on the essel to remoe any liquid
stuck to the end
*otential errors3 inaccurate measurements for reasons gien aboe. What effects
would this hae on the results<
$sing balances
=eer weigh anything directly onto a balance>s pan9 this will contaminate it for
other users. $se a weighing boat or slip of aluminium foil, or paper
=ote reading to 2 decimal places. =.,. When calculating a mean aerage of
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Essential A level Biology Practical Skills
readings, the aerage should also be to 2 decimal places
*otential errors3 samples spilt onto the pan will be measured but not used9 as will
samples left if the weighing boat is not scraped clean. :ow would the results be
affected<
7easuring and Controlling ?emperature
:eating samples3
o Wear safety glasses
o $se a thermostatically4controlled water bath if possible and suitable9 check
the temperature using a thermometer9 do not rely on the temperature
shown on the dial
o =ote that if you are not using a thermostatically controlled water bath,
this would be a good improement you could mention in an ealuation
o /tate the temperature used and the time for which heating is carried out
e.g. ,enedict>s test @ heat for 5-10 minutes in a water bath at 80
o
C
o $se insulation if necessary or possible
*otential errors3 has the temperature aried from the stated alue< :ow would
this affect the results<
7easuring ?ime
$se a stop watch rather than a clock
7ake sure you know which buttons to press before the e%periment startsA
=ote time readings to a suitable number of decimal places. =.,. When calculating
a mean aerage of readings, the aerage should also be to the same number of
decimal places. Could you actually measure the time this accurately<
*otential errors3 how easy is it to know when to start or stop the timer< What
difference would this make to the results<
*reparing Dilutions
/olutions are usually prepared with respect to their
molar concentrations 1molBl or molBdm
3
2 1a mole is a gien number of molecules
of a compound9 1 mole has a mass in grams equal to the relatie molecular mass
of that compound2 or
mass concentrations 1gBl or gBdm
3
2
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Essential A level Biology Practical Skills
,oth these are the amount of a substance per unit olume of solution. i.e.
Concentration . )mount
Colume
Concentration must always be gien units.
5n practicals, you will often be gien stock solutions to use. ?hese are solutions of
known concentration and are aluable when making up a range of solutions of differing
concentrations. ?hey also sae work if the same solution is used oer a long time e.g. a
nutrient solution. /tock solutions are more concentrated than the final requirement and
are diluted as appropriate.
*reparing a dilution series
) series of dilutions is ery useful for a wide range of procedures e.g.
to inestigate changing the concentration of substrate in an en-yme reaction
to prepare calibration cures for colorimetry
How to make a dilution series:
Always start with the most concentrated solution
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7ost
concentrated
solution 1in
e%cess2 1 ml 1 ml 1 ml 1 ml 1 ml
$ndiluted 11D2
1B1D
11D
41
2
1B1DD
11D
42
2
1B1 DDD
11D
43
2
1B1D DDD
11D
4!
2
1B1DD DDD
11D
4#
2
E ml
diluent
Essential A level Biology Practical Skills
1. Decimal dilutions
'ach concentration is one tenth that of the preious one 1log
1D

dilution series2
7easure out the most concentrated solution with a 1DF e%cess
7easure one4tenth of the olume required into a essel containing
nine times as much diluting liquid
7i% thoroughly
+epeat to obtain concentrations 1B1D, 1B1DD, 1B1DDD, etc times the
original
?o calculate the actual concentration of solute multiply by the
appropriate dilution factor
2. Doubling dilutions34
'ach concentration is half that of the preious one
$se twice the olume required of the first, most concentrated
solution
7easure out half of this olume into a essel containing the same
olume of diluting liquid 1e.g. distilled water2
7i% thoroughly
+epeat for as many doubling dilutions as are required
?he concentrations obtained will be G, H, , etc times the original
*otential 'rrors 1+emember thinking about these can help you ealuate your
procedure2
Contamination from syringes9 rinse between use or use new syringes for each
solution to aoid carry4oer of solutions of the wrong concentration
1=ote that when transferring a range of prepared dilutions from sample pots into
test tubes, you should start with the lowest concentration and, if you rinse the
syringe in the ne%t concentration before dispensing it, you can use the same
syringe or pipette2
5naccuracy in measuring olumes9 any slight inaccuracy will lead to compounded
inaccuracies, so the most dilute solutions hae huge errors in concentration 1see
precautions in measuring liquids aboe2
;abel tubes carefully to ensure correct solutions are transferred
7i% solutions before transferring to ensure the correct amount of solent is
added to the ne%t tube
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Essential A level Biology Practical Skills
+ecording Data
Don>t use scraps of paperA
$se a table, which should hae34
?itle
+uled grid lines, '=C;"/5=( );; D)?), 5=C;$D5=( ?:' :')D5=(/
:eadings at tops of columns
?he independent ariable should be in the first column, beginning with the
smallest alue
:eadings should include units. D" ="? *$? $=5?/ 5= ?:' ,"DI "J ?:'
?),;'
/ame number of decimal places for each measurement. ?he number of places
should reflect the accuracy and precision of your measurement. Do not round off
data alues9 this might affect subsequent analysis of your data. 1=, take care if
using the computer as it sometimes automatically changes these2
"bserations of results 1not your interpretation of themA2 Write these in as much
detail as possible
+epeated readings9 use a separate column for each
?hink about any additional columns you may need, and draw them in at the start.
)dditional columns may be used to show step by step calculations e.g. olume
1cm
3
2, time 1s2, 1Btime 11Bs2, rate 1cm
3
Bs2
Calculated mean aerages 1Do not use a greater number of decimal places than you
hae in the raw data2
Design your table to make the recording of data as straightforward as possible,
to minimise the possibility of mistakes
'%plain any unusual data alues in a footnote9 don>t rely on memoryA '.g. forgot to
start stopwatch
Colorimetry
?he spectrophotometer
measures the absorption of
radiation in the isible and
u regions of the
electromagnetic spectrum.
?he spectrophotometer
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Rea
d-
Essential A level Biology Practical Skills
allows precise measurement at a particular waelength. ) colorimeter is simpler, using
filters to measure broader waelength bands 1e.g. green, red or blue light2.
*rinciples of light absorption
?he absorption of light is e%ponentially related to the number of molecules of the
absorbing solute in the solution i.e. KCL, solute concentration
)bsorbance at a particular waelength is often shown as a subscript e.g. )
##D
.
absorbance at ##D nm
?he proportion of light passing through the solution is known as transmittance 1?2
and is calculated as the ratio of the emergent and incident light intensities. 5t is
usually e%pressed as a percentage
?he colorimeter has 2 scales34
o )n e%ponential scale from -ero to infinity, measuring absorbance
o ) linear scale from D @ 1DD, measuring 1per cent2 transmittance
Jor most practical purposes you should use absorbance, which is linearly related
to the solute concentration KCL
'%tension information
Absorbance (A is !i"en by:-
) . log
1D
15
o
B5 2
$sually shown as )
%
, where % . the waelength in nanometres
)lso34
) . l[C]
Where3
= a constant for the absorbing substance 1absorption coefficient2
l . the length of the light path through the absorbing solution
C . concentration in molBl or gBl
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Essential A level Biology Practical Skills
Calibration Cures
,y preparing a set of
standard solutions,
each containing a known
amount or
concentration of a
substance, and then
measuring the
absorbance of each
solution, a Mcalibration
cureN or Mstandard
cureN can be produced. 1=ote that at high concentrations, the relationships aboe do
not hold true, and the straight line relationship shown may become cured.2 ?he line can
be used to estimate concentrations of solute in a test or unknown sample.
:ow to use the colorimeter
1. /witch on
2. )llow 1# minutes for the lamp to warm up and the instrument to stabilise
3. /elect the correct coloured filter 15t is best to use the filter which selects the
range of waelengths most strongly absorbed by the sample because this will gie
the ma%imum reading2. ?he most suitable filter colour is usually the
complementary colour to the solution being tested34
C";"$+ "J /";$?5"= J5;?'+ C";"$+
Ciolet Iellow4green
,lue Iellow
,lue4green +ed
(reen *urple
Iellow4green Ciolet
Iellow ,lue
"range (reen4blue
+ed ,lue4green
*urple4red (reen
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Essential A level Biology Practical Skills
!. 5nsert a reference blank cuette
#. Check the reading is -ero 1-ero if necessary2
&. )nalyse samples
O. Check the scale is -ero at regular interals using a reference blank e.g. after
eery 1D samples
P. Check the reproducibility of the instrument9 measure the absorbance of a single
solution seeral times during analysis. 5t should gie the same alue
5naccuracies may be due to34
5ncorrect use of cuettes
i. Dirt
ii. Jingerprints
iii. ?est solution on the outside of cuettes
Condensation on cold samples 1allow cold samples to equilibrate to room
temperature2
)ir bubbles in samples 1tap gently to remoe2
5nsufficient solution 1refraction of light at meniscus2
Cloudiness of sample 1decant off supernatant to test, after allowing
precipitate to settle2
7icroscopy
*roblems in light microscopy and possible solutions34
#o ima!e$ "ery dark ima!e
microscope not switched on
ob6ectie nosepiece not clicked into place oer a lens
lamp failure
%ma!e blurred and cannot be &ocused
dirty ob6ectie
dirty slide
slide upside down
slide not completely flat on stage
fine focus at end of trael
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Essential A level Biology Practical Skills
'ust and dirt in &ield o& "iew
eyepiece lens dirty
ob6ectie lens dirty
slide dirty
dirty on lamp glass or upper condenser lens
/etting up and using the light microscope
1. /elect low power lens. 7ake sure the lens clicks into position.
2. '%amine prepared slide without the microscope and note position, colour
and rough si-e of specimen.
3. *lace slide on stage, coerslip uppermost, iewing it from the side. *osition
it with stage ad6ustment controls so that the specimen is lit up.
!. Jocus using first the course and then the fine focusing controls. $se both
hands to alter the focusing controls9 this helps keep the controls working
properly and not going out of alignment.
=ote3 ?he image will be reersed and upside down when seen by iewing the
slide directly.
#. Jor higher magnifications, swing in the releant ob6ectie lens carefully
checking there is space for it. )d6ust the focus using the fine control only.
5f the ob6ect is in the centre of the field of iew with %1D ob6ectie, it
should remain in iew with the %!D ob6ectie.
&. When you hae finished using the microscope,
?urn the ob6ectie lens back to %1D
;ower the stage
+emoe the last slide and return to correct section in tray
Clean the stage if necessary
Check eyepiece lenses and ob6ectie lenses are clean
$nplug cable and store tidily
+eplace dust coer
O. (eneral care of the microscope34
=eer force any of the controls
=eer touch any of the glass surfaces with anything other than
clean, dry lens tissue
When moing the microscope, hold the stand aboe the stage with
one hand and rest the base of the stand on your other hand
)lways keep the microscope ertical 1or the eyepiece may fall out2
Do not touch lenses with your fingers
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Essential A level Biology Practical Skills
Do not allow any solent to come in contact with a lens
Do not wear mascara when using the microscope9 it marks the
eyepieceA
'stablishing /cale
7agnification used to obsere a specimen . ob6ectie magnification % eyepiece
magnification.
:oweer, this is fairly meaningless when drawing any image, which could be drawn at any
si-e. Jor this reason, it is essential to add a scale to your drawing. Iou can3
*roide a bar of defined length e.g. 1DD Qm or
(ie an estimated si-e of the ob6ect
7ethods of estimating si-e
1. Compare the si-e of the image to
the diameter of the field of iew34
a. Jocus on the millimetre scale
of a transparent ruler, using
the lowest power ob6ectie
b. 'stimate the diameter of this
field directly
c. $se the information to work
out the field diameters of
higher powers e.g. if the field
at %!D is ! mm, at a
magnification of %1DD will be !DB1DD % ! mm . 1.& mm
2. (reater accuracy can be obtained if an eyepiece graticule 1micrometer2 is used34
a. (raticule carries a fine scale and fits inside an eyepiece lens
b. (raticule is calibrated using a stage micrometer i.e. a slide with a fine scale
in it 1Iou will do this yourself2.
c. "nce you calibrate your eyepiece graticule for each ob6ectie lens used, you
can use it to measure ob6ects
d. 5n the e%ample below, the scale reading is multiplied by 2.&# Qm to gie the
alue in micrometres.
e. )oid putting too many significant figures in any estimates of dimensions9
there may be quite large errors inoled
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Essential A level Biology Practical Skills
(ecordin! by biolo!ical drawin!s
?he making of accurate drawings is an essential skill for )danced leel biology
students. /tudents are required to make drawings of e%ternal features of whole
specimens, parts of specimens, dissections and prepared slides.
Drawings must be done on white )a)er usin! shar) )encil. /tudents should note the
following points3
1. ) drawing should be genuine and accurate record of what has been seen. Do not
copy diagrams from books with little resemblance to the specimen, dissection or
prepared slide under inestigation.
2. Jor low power drawings, only the outline of the structures or tissues needs to be
drawn. 5ndiidual cells are required only for high power drawings.
3. Draw with bold, dark, smooth lines. /hading and the use of crayons are not
encouraged.
!. )oid making drawings on both sides of a paper.
#. 5n making a drawing, first decide what you want to show. ?hen plan your drawing
to fit on the page. 5t is important that the arious parts of the structures are
drawn in proportion.
&. Drawings should be large, neat and tidy.
O. )ll releant structures should be fully and clearly labelled usin! )encil* 'ach
label should be connected to the appropriate part of the drawing by a clear
labelling line. ?he labelling lines should run as hori-ontal as possible and should not
intersect with one another. Do not label too close to the drawings, and do not
write on the drawing itself. Distribute the labels well around the drawing so that
the labelling lines can be kept reasonably short.
P. 'ery drawing should include a title, the magnification power, and the plan of iew
of the specimen 1such as transerse section, longitudinal section2.
E. 7ake additional drawings if important details are too small to be shown in a low
power drawing. ?his can be done by making a simple drawing of the main features,
and other drawings on details of small parts only. Jor e%ample, the recording of a
transerse section of a stem should include a low power plan of the section and
high power drawings of different cells types as required showing the detail of the
cells.
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Essential A level Biology Practical Skills
1D. 5f suffices to show the internal structural details in a small representatie sector
of the specimen which shows repetitie features.
11. 5t is sometimes appropriate to make short succinct notes close to the labels.
/uch annotated drawings are particularly aluable as they combine a record of
structures with related functions andBor biological interests. Iou may want to
label an organelle to describe staining e.g. nucleus 1darkly stained29 starch grain
1blue4black2
'rawin! +ra)hs
(raphs gie a isual impression of the content and meaning of your results. ?ables
proide an accurate numerical record of data alues.
(raphs should34
o include all material necessary to coney the appropriate message without
reference to the te%t
When drawing graphs34
o Collect all data alues to be plotted
o Consider whether a graph is the best way to present the data
o Choose a concise e%planatory title to establish the content
o Consider the layout and scale of the a%es carefully
o $se the % a%is for the independent ariable
o $se the y a%is for the dependent ariable
o When neither ariable is determined by the other, or where ariables are
interdependent, the a%es may be plotted either way round
o 'ach a%is should hae a descriptie label showing what is represented and
including units of measurement, separated by B or written in brackets.
o 'ach a%is must hae an appropriate marked scale, showing the location of all
numbers used. Jill the aailable space on the paper
o 5f scale breaks are used, show them clearly
o Choose the symbols for each set of data points, if plotting more than one
set of data
o 5nclude a key to symbols of different data sets if necessary
o ?o plot ery large or small numbers, the plotted alues may be measured
numbers multiplied by a power of ten e.g. 1D
43
% population of bacteria B mm
3
o Draw a trend line for each set of points3
Cure<
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Essential A level Biology Practical Skills
/traight line<
*oints 6oined with a ruler< 1?his is really only alid if there is -ero
error. i.e. all the plotted points are precise and accurate2
o )lways draw the simplest line that fits the data reasonably well and is
biologically reasonable.
o ?ry to aoid the need to e%trapolate plotted cures by better e%perimental
design. 1?his could be a point for ealuation of an e%periment2.
o =eer allow a computer programme to dictate what a graph looks like9 make
sure you can alter scales, labels, a%es etc and make appropriate selections.
Draw cures freehand if necessary.
%nter)retin! +ra)hs , Analysis and -"aluation
1. Describe the relationship between the ariables, quoting data.
2. '%plain what the relationship means with reference to the biological principles
inoled.
3. Consider the content3
a. What was the aim B hypothesis of the inestigation<
b. Why were the obserations made<
!. Consider what kind of graph is presented9 is it an appropriate choice for the data<
#. ;ook carefully at the scale of each a%is.
a. What is the starting point<
b. What is the highest alue<
c. Do the alues start at -ero< ) non4-ero a%is emphasi-es the differences in
measurements by reducing the range of alues coered by the a%is.
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Essential A level Biology Practical Skills
&. '%amine symbols and trend lines9 e.g. if 2 conditions hae been obsered while a
ariable is altered, when do they differ from each other9 by how much and for
how long<
O. 'aluate errors34
a. ;ook for ariability in the data9 hae anomalies been recogni-ed< :ae they
been included in the trend line< 5s it reasonable to ignore such points<
b. 5f mean alues are presented, the underlying errors could be large
Page 15 of 16
Essential A level Biology Practical Skills
c. :as a graph been e%trapolated correctly< ?here can be no guarantee that
relationships will hold under new conditions, so the e%trapolation may be
inaccurate.
d. 5s the trend line appropriate< Would a cure B straight line be more
appropriate< Consider the theoretical alidity of the line. ) straight line
implies one factor aries in direct relationship to another9 the true
situation may be more comple% e.g. an e%ponential relationship could be
more correct.
Page 16 of 16