FACULTY OF ENGINEERING & LIFE SCIENCES
MGS1133 BASIC BIOCHEMISTRY
LABORATORY REPORT
(EXERCISE 1.0: USING PIPETTORS)
LECTURER: DR. HAMIDAH ABU BAKAR
NAMES OF STUDENTS AND MATRIC NUMBERS:
 MUHAMAD SYAZWAN BIN MISRAN 4174015421
 MOHAMOUD MOHAMED OSMAN
4173010591
 SAFIA MOHAMED ABDALLAH
4171003981
 NADIRA ABDIQANI ADEN
4171008441
 HAMDA HUSSEIN OSMAN
4173009881
INTRODUCTION
In Biochemistry, pipettes are used in nearly all experiments. In order to obtain accurate and
precise data, correct operation of the pipettes is imperative. For this reason, the course is started
with an exercise to familiarize everyone with correct pipetting techniques.
A microliter pipettor is a variable-stroke piston pipette. The volume indicator consists of three
number dials and is read from top to bottom. The three digits indicate the volume selected. The
black digits on the P-20 and P-200 show microliters and the red digits on the P-20 show tenths of
microliters. For the P-1000, the digits in red represent milliliters and the digits in black represent
microliters. Ranges out of these are not used.
Accuracy is the closeness to which the dispensed volume approximates the true volume as set on
the pipette. Accuracy is expressed as mean error or % error, the percent by which the mean value
of a large number of replicate measurements of the same volume will deviate from the expected
or “true” volume. The accuracy of these pipettes is determined by the factory calibration and
checked gravimetrically using distilled water and an analytical balance. Careful use will maintain
this calibration and accuracy throughout the semester.
Precision refers to the “scatter” of individual measurements around the mean of replicate
measurements. It can be expressed as sample standard deviation.
OBJECTIVES
1. To learn how to use pipettors.
2. To be able calibrate the pipettors.
MATERIALS
1. Micropipettors and tips
2. Volumetric pipet (10 mL)
3. Measuring pipet (graduated pipet)
4. Pipette filler
5. Distilled water
6. Weighing boat
7. Balance
PROCEDURE
Operation of the Microliter Pipette
1.
The volume is set by turning the volume adjustment knob at the end of the pipette until the
correct volume shows on the indicator.
A new disposable tip is attached to the pipette shaft. It is pressed firmly with a slight twisting
motion. The correct size of tips are used for each pipette.
When gathering sample, the plunger is pressed to the first stop. This part of the stroke is the
calibrated volume displayed on the digital volume indicator. The plunger cannot be pressed all
the way down, or the solution will be drawn up too much.
2.
The microliter pipettor is held vertically, immersing part of the disposable tip into the
sample.
3.
The push-button is allowed to return slowly to the up position. It cannot be snapped up. This
is done slowly and the tip is kept submerged in the solution to prevent any air bubbles from
entering the tip that will mess up the volume measurement.
4.
A few seconds is waited to ensure that the full volume of sample is drawn into the tip.
5.
The tip is withdrawn from the sample liquid. Any liquid that remained on the outside of the
tip is wiped off carefully with a lint-free tissue, while not touching the orifice.
6.
The sample is dispensed by touching the tip end to the sidewall of the receiving vessel and
the plunger is depressed slowly to the first stop. Two seconds is waited. Then the plunger is
pressed to the second stop (the bottom stroke), expelling any residual liquid in the tip.
7.
With the plunger fully depressed, the microliter pipettor is withdrawn from the vessel
carefully, with the tip sliding along the wall of the vessel.
8.
The plunger is allowed to return slowly to the up position.
9.
The tip is discarded. Different tip is used each time different material is gathered/dispensed
so that the concentrations of solutions are accurate.
Analysis
1. Suitable pipet is acquired with the correct volume size required as in the Table 1.
2. Weighing boat is placed on the balance and the weight is tarred to zero.
3. The designated volume of distilled water is drawn up and dispensed onto the weighing
boat. The weight of the water is recorded. The procedure is repeated twice.
4. The result is analysed.
RESULTS
Table 1
Designated
Volume (mL)
Weight
(g)
Type of Pipet
0.05
0.05 g
Microliter pipettor
P100
0.2
0.2 g
Microliter pipettor
P1000
0.015
0.015 g
Microliter pipettor
P20
0.105
0.105 g
Microliter pipettor
P20
2.35
2.35 g
4.02
4.02 g
Pipette 10ml x
0.1ml
Pipette 10ml x
0.1ml
Analysis
Conversion to µL:
0.05mL x 1000 = 50 µL
Weight of the water = 0.0498 g
Conversion to µL:
0.2mL x 1000 = 200µL
Weight of the water = 0.2058 g
Conversion to µL:
0.015mL x 1000 = 15µL
Weight of the water = 0.0005 g
Conversion to µL:
0.105mL x 1000 = 105µL
Weight of the water = 0.098 g
Weight of the water = 2.25 g
Weight of the water = 3.9223 g
DISCUSSION
1.
Based on the test that was made, the accuracy is calculated as below :
Designated Volume
(mL)
0.05
0.2
0.015
0.105
2.35
4.02
2.
Weight of the distilled water (g)
Accuracy (%)
0.0498 g
0.2058 g
0.0005 g
0.098 g
2.25 g
3.9223 g
99.6
97.1
3.33
93.33
95.74
97.57
In order to get an accurate result, suitable range of pipette (or micropipette) is used. For
example, it is inaccurate if P100 micropipette is used for 0.105ml of distilled water, but it
is more accurate if P20 micropipette is used.
3.
For the method of drawing up distilled water into the micropipette, doing it too fast will
cause inaccuracy. Drawing up distilled water is done slowly in order to gain accuracy.
4.
The plunger is pressed only to the first stop and not to the second stop before filling the
micropipette tip. Pressing all the way down will cause too much of distilled water being
withdrawn.
5.
Different tips is used for different sample in order to avoid inaccuracy.
CONCLUSION
The accuracy in using the pipettors is determined by the correct way of using the pipettors, such
as using suitable range of pipettor, drawing up sample slowly, not pressing the plunger all the
way down before filling up the micropipette tip, and using different tip for different sample.
REFERENCES:
Title: Mastering the micropipette. Accessed from: http://capricorn.bc.edu/bi204/wpcontent/uploads/2013/08/3-micropipettes.pdf. Accessed date: 30/10/2017.