# Calibration of Pipets

A Statistical Point of View

Types of Pipets
Two Main Types
- TC (to contain) - TD (to deliver)

Purpose of Calibration
• To determine the random error introduced by human error. Then to determine systematic error introduced due to poor manufacturing tolerances in either the balance or the pipet itself.

by transferring aliquots of known density from the pipet to a weighing vessel and measuring the masses transferred. .Gravimetric Titration • Conventional volumetric calibration procedure using gravimetry i.e.

• Experimental conditions used in calibration should match as closely as possible the conditions of the experiment itself .Gravimetric Calibrations • Gravimetric calibration uses an assumed correct and working balance to give a particular amount of a fluid at a known weight using a standard such a H2O.

effective.Advantage to Gravimetric Titration • • • • • Easy to set up Distilled H2O is easy to acquire Cheap Can be done with very few aliquots Calibration of volumetric devices by gravimetry is simple. and wellknown and under ordinary circumstances this procedure and associated data analysis are straight forward .

Disadvantages to Gravimetric Calibration • Not an accurate procedure • Assumes low error and a precisely calibrated balance • The balance MUST be properly calibrated • To improve accuracy. many aliquots must be taken • Taring can be time consuming depending on vessel .

.Analysis • Plot large amounts of aliquots and see if they drift aimlessly about the mean. • The smaller the aliquot. To counteract this taring is best done performed as well as containment of evaporation along with large numbers of aliquots measured. if they don’t then some other systematic error must be found and corrected. the larger and easier errors are to introduce.

EXPERIMENTAL PROCEDURES • • Two procedures: tared and untared Same equipment and materials used for both procedures .

0% capacity Mean: 0. the nominal volume delivered by the micropipette was 0.0005 g.0084 mL The temperature of the water was measured to be 21. .2 degrees.0.1906 g and a standard error of 0.00051 g System Correction: -0. With a mean of 0.7 degrees Celsius +/.9978 g/mL.1911 g.EXPERIMENT 1 (TARED) – Amount of trials: 40 aliquots • • • • • Each aliquot: 20.19069 g Standard deviation: 0. and the density of the water was calculated to be 0.0032 g Standard error: 0.

standard deviation. . and standard error were calculated through extrapolation of a residual plot trend.0006 g • System Correction: -0.0034 g • Standard error: 0.19064 g • Standard deviation: 0.EXPERIMENT 2 (UNTARED) • Amount of trials: 40 aliquots • Each aliquot: 20.0094 mL Values for mean.0% capacity • Mean: 0.

0005 g System Correction: -0. the difference in system correction was 0.19064 g • Standard deviation: 0. Evaluating these values with a t test affirms that these values are consistent.0032 g Standard error: 0.19069 g Standard deviation: 0.0010 mL and the standard error estimate was 0.0006 g • System Correction: 0.0094 mL Mean: 0.0034 g • Standard error: 0.0008 mL.COMPARISON OF EXPERIMENT 1 AND EXPERIMENT 2 • • • • Experiment 1 Experiment 2 • Mean: 0.0084 mL Factoring in the standard deviation and standard error. .

Results Experiment One vs. Experiment Two vs. Lab1 (Calibration of glassware) .

• Standard deviation. tared immediately before each weighing .Experiment One • Weighed 40 aliquots .. .0005 being rate of water evaporation * time interval.0084 +.0032g • Volume change of -..0005 • The .

Experiment One (Cont.) The independent random component can be found by the residual variable being subtracted from the previous residual. .

) • Volumetric Variance .02x10^-5 ml^2 based on 39 degrees of freedom which leads to a volumetric standard deviation of Sv=.0032 mL based on 39 degrees .Sѵ₂= 1.Experiment One (Cont.

0006 being rate of water evaporation * time interval...Experiment Two • Weighed 40 aliquots . • Standard deviation.00094 +. .0006 • The . Did not tare before each weighing .0034 • Volume change of-.

) .Experiment Two (Cont.

16x10^-5 ml^2 based on 37 degrees of freedom which leads to a volumetric standard deviation of Sv=.Experiment Two (Cont.0034 mL based on 37 degrees.) • Volumetric Variance . .Sѵ₂= 1.

30ml . 40ml and 50ml of distilled water. • Measured mass of water • From mass and density we found the volume . 20ml .Lab 1 (Calibration of Glassware) • Used measurements of 10ml . .

Lab One ( cont.) Graphed experimental amount of water vs. the theoretical amount of water. .

Discussion • The difference of change in volume between the 2 experiments is 0.0008 mL. • A (t) test for the hypothesis that the difference is within statistical scatter of zero – which leads to an acceptance of the hypothesis at any reasonable confidence interval.0010 mL and the standard error estimate of the difference is . .

Discussion (Cont.) • For the volumetric variances we take the ratio (1.14 – which is less than the tabulated F.16x10^-5)mL / (1.statistic for any reasonable confidence interval .04x10^-5) mL • The ratio of the volumetric variance estimates gives us 1.

0034 mL are consistent.Discussion (Cont. .) • Thus we may accept the null hypothesis and the two variances are from the statistical population and conclude that the volumetric standard deviations of .0032 and .

7 . Assoc. 1964 . Statist. Boston.dept of chemistry .com. J. Anal.J the statistical Analysis of Experimental Data . Schwartz).Works Cited • Calibration of pipettes – A statistical view (Lowell M. University of Massachusetts . 1957 .J Am. 552-566 • Mandel . S. 1989 61. 1080-1083 • Meyer . J. 12. 52 . Willey : New York 1975 chapter 33 • Mandel . L Data for Analysis for scientist and Engineers . Willey . New York.8 . Sections 12. Chem . ( Via ACS.

Resource and Discussion • Statistical uncertainties and accuracies – Matching experimental conditions – Properly calibrated weighing machine • The pipet maybe used to contain or deliver the aliquot • Two Ways – Calibrated series of vessel (tared) – Using single vessel (Untered) .

Precision of the Experiment • Highly reliable data – 30 times or more replications – Use large number of vessel OR – Use one vessel to transfer series of aliquots .

Tared Vs Untared Tared • Taring vessel is time consuming and troublesome • Evaporation of liquid negligible • Data analysis is simpler Untared • No need to tare the vessel each time • Must account evaporation of liquid • Data analysis very cumbersome .

Two Errors • Systematic error – Denote by ∆v – Partly comes from the manufacture tolerance – Partly result from the fluctuation of dispensing technique between manufacture and analyst • Random error: – Denote Єv – Result from successive aliquot delivered – Magnitude depending on the operation sill of the analyst – Taking weighing data of the aliquot .

• Balance must be able to operate with minimal mechanical and electrical fluctuation .Points of Interest • The density of the liquid be known • The liquid must be equilibrated at STP condition • When plotting the data. make sure the weighing data scattered randomly about the mean.

Condition STP 5. Statistical replications 30 times or more Calibration of glassware 1.Comparison with Our 1st Lab Calibration of Pipet 1. Condition STP 5. Used only one vessel 4. Based on volumetric calculation 3. Same 2. Statistical replications only five times . Uses Single or series of vessel 4. same 3. Assumed the balance is calibrated 2.

Data analysis is less complicated and straight forward . Data analysis is time consuming and complicated Calibration of glassware 7. Calibration of mL glassware's (difference 1000 times) 9. Calibration of Micro pipet (1 micro liters) 9. Evaporation neglected 8.Compression Continued Calibration of pipets 7. Evaporation considered 8.

Conclusion .

and all systematic errors are taken into account.Error Reduction • Calibration of pipets is a delicate procedure where much care must be taken to assure as little human error as possible is introduced. .

. lost cost/high value procedure to ensure measurement precision.Effectiveness • Gravimetric titration is an easy to perform.

.Availibility • Almost universally available due to all equipment being easy to procure. distilled water. IE balance. and pipets.

Reliability • Can be very reliable when care is taken and large amounts of aliquots are used to reduce the human error introduced. Reliability is low with less aliquots measured. . Also assumes working balance.

Questions .