Post Lab #2 Pipette Calibration PDF

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CHEM 3214-0A1 Pipette Calibration Post Lab 09/13/17

Introduction The purpose of this laboratory experiment is to exercise pipette calibration and gain familiarity on the proper usage of laboratory pipettes. Pipettes are useful during laboratory experiments due to an important feature in that they offer an accurate and precise method of transferring desired volumes of liquid. There are three types of pipettes most commonly found in a laboratory setting which are volumetric, measuring, and mechanical micropipettes. In order to effectively take advantage of instruments that offer accurate and precise measurements, proper usage must be enforced as well as frequent calibration. Following proper standard operating procedures on the usage of pipettes will greatly increase the accuracy and precision of each measurement. The relative error of the user will decrease as well as a reduced standard deviation will be observed. Certain measures must be enforced such as keeping devices clean and dry prior to each use. Improper use of laboratory equipment will result in a higher standard deviation. Frequent calibration of laboratory equipment enables frequent inspection on the functionality of the instrument at question. During a pipette calibration procedure, the mechanical parts that enable the instrument to perform properly are examined. This ensures that the quantities that are being measured are accurate to maintain the integrity of the data being collected. When the mechanical parts of a measuring instrument have been compromised, it is likely that the accuracy of the instrument will deviate away from the accepted value at a higher probability. To conduct the pipette calibration, aliquots of water are repeatedly pipetted and weighted. A mean and standard deviation is calculated which are utilized to calculate the coefficient of variation (CV). The coefficient of variation takes the standard deviation and divides it by the mean and multiplied by 100. This describes the amount of variability compared to the mean which gives insight on the precision and accuracy of the pipette being calibrated. A CV is acceptable if it is less than or equal to

±

3%. A CV greater than ± 3% is not acceptable

and cannot be used in collecting parametric data. A CV greater than

± 3% indicates that the

pipette exhibits mechanical failure that interferes with the precision and accuracy of the measuring device. To achieve parametric data that best suits the actual measurement calibration, must be conducted frequently.

Materials 

Analytical balance



Micropipette



Beaker



DI water

Hazards and Safety (MSDS) Chemical Name

Chemical Formula

Molecular Mass (g/mol)

Melting Point Boiling Point (C°) (C°)

Hazards

Water

H 2O

18.02

0

Nonhazardous

100

Methods/Procedure 1. Place small plastic weighing boat on analytical balance. 2. Depress pipette plunger until resistance is expressed. 3. Place the tip of the pipette into beaker containing water. 4. Slowly release plunger until fully extended to collect aliquot of water. 5. Remove excess water on pipette tip. 6. Deliver the water to the weighing boat by fully depressing the plunger on the micropipette. 7. Record the weight of the water delivered on the weighing boat on a pipette calibration form. 8. Using the same pipette tip repeat procedure of weighing water aliquots 5 times for 3 trials. 9. Calculate the CV following the formula a. CV= Standard Deviation in Volume/Average Volume x 100

Results and Data

Table 1. Mass of Water Aliquots at Varied Volumes Trial

Expected Volume (µL)

Mass (g)

Calculated Volume (µL)

55 µL Water Aliquots 1

55µL

0.0556g

55.6 µL

2

55µL

0.0524g

52.4 µL

3

55µL

0.0557g

55.7 µL

4

55µL

0.0551g

55.1 µL

5

55µL

0.0521g

52.1 µL

60 µL Water Aliquots 1

60µL

0.0585g

58.5 µL

2

60µL

0.0607g

60.7 µL

3

60µL

0.0625g

62.5 µL

4

60µL

0.0601g

60.1 µL

5

60µL

0.0604g

60.4 µL

102 µL Water Aliquots 1

102µ

0.1005g

100.5 µL

2

102µ

0.1020g

102 µL

3

102µ

0.1022g

102.2 µL

4

102µ

0.1016g

101.6 µL

5

102µ

0.1006g

100.6 µL

volume (1 µL trial 1) =

=

0.0556 g 0.001

mass volume

= 55.6µL

Table 2. Parametric Approach to Analyzing the Data Data Parameters

55µL

60µL

102µL

Mean x (n=5)

0.0542

0.060

0.101

Standard deviation (s)

0.002

0.0014

7.89x10-4

Coefficient of Variation (CV)

3.69

2.33

0.781

Is CV acceptablea ?

No

Yes

Yes

a

CV is acceptable if, CV ≤ ± 3 % Mean x (55 µL) = =

1 n ∑x n i=1 i 0.0556 + 0.0524 + 0.0557 + 0.0551 + 0.0521 5

Standard deviation (s) (55 µL)

= 0.0542

=

√

√

∑ (x i−x )2

=

n−1

(0.0556−0.0542)2 +( 0.0524− 0.0542)2 +( 0.0557−0.0542 )2 +(0.0551 −0.0542)2 +(0.0521−0 5−1 = 0.002

Coefficient of Variation (55 µL) s = 100 × x = 100 ×

0.002 = 3.69 0.0542

Analysis/Discussion and Conclusion

The purpose of this laboratory experiment was to exercise pipette calibration and gain familiarity on the proper usage of laboratory pipettes. For this experiment aliquots of 55,60 and 102 µL water were measured for pipette calibration. For the 55µL of water, a CV higher than 3% was calculated. In addition, this trial had the largest standard deviation. This could have been caused by improper usage of the pipette. The most accurate measurements were from 102µL giving a relatively small standard deviation and CV making it acceptable. The reason why the first trial gave such a large standard deviation is because it was not properly calibrated, and for the third trial a smaller standard deviation was calculated making the pipette now calibrated.

Having laboratory equipment that is well calibrated is important because it can give precise and accurate measurements. In addition, it maintains the integrity of the data that is being collected and relative error is reduced. Large standard deviations signify improper use or uncalibrated instruments. Coefficient of variation plays an important calculation that could be ruined for improper calibration of an instrument because it gives an idea about the precision and accuracy of the instrument being calibrated. Instruments improperly calibrated can cause unprecise and inaccurate data in an experiment. There can definitively be human error when using laboratory instruments. If someone does not know how to properly use an instrument it can cause unprecise and inaccurate data. Knowledge of usage of instrumentation is always important to keep data integrity. When depressing the pipette twice, the measured volume will not be affected because when depressing it twice you get all the material “out”. Sometimes when working with small quantities like microliters is hard to get all the material out of the pipette since is such a small quantity. That may be another reason why the standard deviation for the first trial was so large. This can be a positive thing since you are getting a more accurate measurement by getting all the material in the beaker. In conclusion, calibration is general with any instrument is an important job since it maintains accuracy and precision in measurements when conducting an experiment. Improper calibration of instruments can result of lack of precision and accuracy along with large standard deviations....