Micropipettes

Micropipettes are the standard laboratory equipment used

to measure and transfer small volumes of liquid

Used extensively in laboratory for

Accurate

Precise measurements
 Types of micropipette

1. Fixed volume

Those which are designed and supplied by manufacturer to

dispense a specific, fixed volume of liquid. This volume
cannot be altered.

2. Variable volume

Those in which the user can adjust the volume of liquid to

be dispensed over a range specified by the manufacturer.
Parts of micropipette
 Sizes of micropipettes
Micropipettes in this laboratory come in three different sizes

 P20 0.5-20μl
 P200 20-200μl
 P1000 100-1000μl
 Operating micropipette
Set the volume
The volume adjustment dial near the top of the
micropipette
Reading volume
 Pipette Tips

There are two sizes of tips

 Small Yellow for P20 and P200
 Large Blue for P1000
Inserting the Tip
 Select the correct size tips.
 Open the box without touching the tips with your hands.
 Insert the micropipette shaft into the tip and press down
firmly. This will attach the tip to the shaft.
 Remove the micropipette with the tip attached
 Plunger stopping

The plunger will stop at two different positions

 First stopping points is the point of initial resistance and

is the level of depression that will result in the desired

volume of solution being transferred
 The second stopping point is when the plunger is

depressed beyond the initial resistance until it is in

contact with the body of the pipette
 This second stopping point is only used for the
complete discharging of solutions from the tip
 Micropipette Rules

Never adjust the volume beyond the range of the

micropipette
Never force the volume adjustor dial

Do not drop pipettes

Always use a smooth motion when using the pipettes

Always keep pipettes upright

Never lay a pipette on the bench top

Always choose to appropriate size pipette for the volume

you are measuring.

 Calibration of Micropipette

Calibration of pipette is carried out by gravimetric

method
The volume delivered by the pipettes is determined by

weighing the amount delivered and dividing this by the

density of water

Principle
Under a constant temperature and atmospheric pressure,

the density of distilled water is constant. The volume of

water can be determined by weighting dispensed water.
Factors affect accuracy of measurement
 Ambient temperature
 Atmospheric pressure

 Relative humidity
These factors are usually combined to give the Z factor,

used in calculation of volume of water

Calibration of pipette is determine by

 Accuracy

 Precision
 Accuracy and Precision
Accuracy means the closeness with which the dispensed

volume approximates the volume set on the pipette

Accuracy is specified as mean error, the average deviation of

replicate measurements from the expected set volume

Precision is the "scatter" or reproducibility of individual

measurements of the same volume

Precision can also be expressed as standard deviation OR

coefficient of variation
Accuracy and Precision

Relative accuracies error are generally about 1% or less

Precision is less than 1 % except when transferring the

smallest recommended volume for a given pipette model

Using the pipettes to transfer volumes which are below

the recommended range will introduce larger errors

 Material and equipment
 Pipette and tips
 beaker
 Distilled water
 Temperature meter
 Analytical balance
 Atmospheric pressure meter
Procedure
 Measure distil water using the pipette of interest (10x)

 Weight each measurements using analytical balance

 Calculate mean, CV and accuracy error

 Calculation
Convert the weight unit of measured value into the volume

unit of measured value using the following formula:

 Volume (ml or ul) = Weight (mg or ug) x Z
Z value: conversion factor, which is conversion of density

Calculate the average (Mean), accuracy, standard deviation

(S.D.) and imprecision (C.V.)

C.V =

Accuracy =
Example
Using 50ul of pipette we measure distilled water and the

weight of water gives the following result. calculate presion

and accuracy
 0.048 mg
 0.045 mg
 0.051 mg
 0.049 mg
 0.053 mg
Calculations for Precision and Accuracy

Arithmetic mean
x
 x
N
Standard deviation

Coefficient of variation

%CV  SD 100
x
 Result
Questions Answers
1. What is the mean? 0.0492 mg
2. What is the coefficient of 6.1%
variation (%C.V.)?

3. What is the standard deviation? 0.003 mg

4. What is the accuracy error of this [(.05-.0492)/.05] x 100 =
pipette? 1.6%

5. Why or why not is this pipette Not acceptable since

acceptable to use? accuracy error > 1% and
precision > 2%
End of chapter two