What You Need to Know About Pipette Calibration

A pipette is a laboratory instrument used in medical and chemistry laboratories to measure and transfer small volumes of liquid for tests or experiments that demand high accuracy. Pipettes come in various types from simple fixed-volume pipettes to more advanced adjustable, multichannel or electronic pipettes. They can dispel volumes of liquids from 10 ml or greater to micro-volumes of 1 ul or less.

Pipette Calibration

Component integrity is critical to the performance of a pipette. There are a number of internal factors that can influence the calibration accuracy including internal friction, worn sealing mechanisms, damaged tips, a corroded piston or instrument cleanliness.

Regular pipette calibration is an integral part of good laboratory practice (GLP) and reduces waste and rework due to errors. Pipette calibration is necessary to maintain accurate results, performance, and longevity of the pipette.

Calibration intervals vary depending on how often the pipette is used and the types of liquids that are dispensed, but to ensure consistent repeatability, calibration should be performed at a minimum of every six months.

Pipette Calibration Theory

At a temperature of 68°F (20°C) and one standard atmosphere of pressure, the density of water is a known constant of 1 g/mL. The volume of water can then be determined by weighing a sample of dispensed water in an accurate analytical balance.

Variations in temperature, humidity, and atmospheric pressure will have an influence on the accuracy of the measurements. These factors are combined to calculate the ‘Z factor’, which is a conversion factor based on the density of the water. The Z-factor is applied in the calculation of the water volume to determine the mass of the dispensed sample and thus the accuracy of the pipette.

Pipette Calibration Procedure

The most common method of pipette calibration is by gravimetric analysis, which involves dispensing samples of distilled water into a container and the weight of the water measured by a precision analytical balance. Gravimetric analysis is preferred by most laboratories due to its simplicity and the traceability of the distilled water as an absolute standard.
Temperature changes in the testing environment or equipment may significantly impact the calibration, so it is important to ensure that the distilled water and pipette are placed in the calibration environment for at least an hour before starting the calibration so the equipment can adjust to the temperature. Precautions should be taken to ensure that the environment is kept as constant as possible during the calibration.

The general procedure for the calibration of pipettes is:
  1. Measure and record the temperature of the distilled water in the beaker.
  2. Place a small beaker or container on the analytical balance platform. Tare the beaker or container weight.
  3. Select the proper tip for the appropriate measurement volume and ensure the tip is clean. Dispense the selected water volume three times to unclog the tip and remove any bubbles in the pipette.
  4. Dispense the water slowly into the weighing container. Record the weight indicated on the analytical balance. Remove the water, clean the weighing container and repeat the process ten times.
  5. Calculate the volume dispensed by the pipette by using the equation ‘V = W x Z’ where V is calculated volume of dispensed water, W is the recorded weight of the distilled water test sample and Z is the Z factor. Average the values obtained from the ten tests.
  6. To finish the calculation, the equation ‘A = 100 x Vav/V0 is used, where A is the accuracy.

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