| Operation and Calibration of Micropipette |
1.0 OBJECTIVE To lay down a Procedure for operation and calibration of micropipette in microbiology department. 2.0 SCOPE This procedure is applicable for operating micropipette used for pipetting liquid in accurate quantities in smaller amount for BET testing as well as microbiology limit test and Microbial culturing in the Quality Control Department of Microbiology Area. 3.0 RESPONSIBILITY • Microbiologist: is responsible for carrying out the established procedure • Head of Microbiology : is responsible for ensuring that the SOP is followed correctly. 4.0 ACCOUNTABILITY • Department Head 5.0 PROCEDURE 5.1 General instruction 5.1.1 The Micropipette must be handled with great care to avid damage. 5.1.2 A damaged micropipette may produce imprecise measurement and this could affect the results. 5.1.3 Do not adjust the volume of the micropipette above or below the recommended range. 5.1.4 When not in use, lay the micropipette flat on the workbench or hang it on the pipette stand. 5.1.5 Never use the micropipette without a tip attached. 5.1.6 Do not keep the pipettes in horizontal position when a filled pipette tip is attached as this may results in liquid entering the pipette. 5.1.7 Use micropipette only for aqueous solution and clean the micropipette after use. 5.2 Operating procedure 5.2.1 The delivery volume is set using the operating button on the top of the micro-pipette. 5.2.2 To increase the delivery volume, turn the operating button clockwise and decrease turn it anticlockwise. 5.2.3 Ensure that the desired delivery volume clicks into the place and that the digest for the volume are completely visible on the handle display. 5.2.4 Hold the micropipette in one hand and with the other hand turn the volume adjustment knob until the volume indicator display the desired volume. 5.2.5 Hold the micropipette so that the tip ejector button is facing the user and the thumb is free to depress the push button. 5.2.6 Select and open a box containing the correct size tips to be used. Or unwrapped Depyrogenated tips. 5.2.7 Place the tip on the shaft of the pipette. Press tip firmly using slight twisting action to ensure air tight sealing. 5.2.8 Hold the pipette in nearly vertical position. Using the thumb, depress the push button to the first stop to micropipette. 5.2.9 Immerse the micropipette tip about half-way into the reagent which is to be measured. 5.2.10 Make sure the reagent solution does not touch the micropipette shaft. 5.2.11 With the pipette tip immersed in the reagent solution, draw the reagent into the tip by slowly raising the thumb and releasing the push button. 5.2.12 Remove the tip from the reagent solution by lifting the micropipette. 5.2.13 Visually examine the tip to make sure it contains reagent solution and there are no bubbles present. 5.2.14 Move the micropipette towards the transfer container which is to receive the measured reagent. 5.2.15 Expel the reagent inside the tip into the transfer container by placing the tip in the transfer container and slowly depressing the push button with the thumb to the first stop. 5.2.16 Wait until liquid stops flowing and press down the push button to the second stop until the tip is completely empty. 5.2.17 With the thumb still holding down the depressed push button, lift the micropipette and tip out of the transfer container. 5.2.18 With the micropipette and tip put out of the transfer container, slowly raise the thumb to release the push button. 5.2.19 Discard the plastic tip. 5.2.20 Place the pipette back into the respective stands after use. To Read this SOP Click Here SOP for Entry and Exit in Microbiology Testing Area. 5.3 Calibration 5.3.1 Micropipettes are calibrated using purified water under constant room temperature 25°c. 5.3.2 Check and ensure that the analytical balance is calibrated before use. 5.3.3 Weight the glass beaker on the calibrated analytical balance and “TARE” the weight of beaker. 5.3.4 Fix the tip and deliver a fixed volume of purified water into the tarred beaker using the pipette to be calibrated. Noted down the weight and tare the weight of the beaker in micro. 5.3.5 Repeat the procedure using 10 deliveries in the same beaker and note down the weight during each delivery. 5.3.6 Ensure that the temperature is maintained at 25°± 1°c during the repeated measurement. 5.3.7 Working range and the calibration points are, |
| Micropipette Range (mL) | Calibrate at volume (mL) | Maximum Permissible Errors (ISO 8655) | ||
| systematic error (mL) | Lower Limit | Upper Limit | ||
| 20 – 200 | 20 | ± 0.80 | 19.2 | 20.8 |
| 100 – 1000 | 50 | ± 0.80 | 49.2 | 50.8 |
| 200-1000 | 100 | ± 1.60 | 98.4 | 101.6 |
| 200 | ± 1.60 | 198.4 | 201.6 | |
| 500 | ± 8.0 | 492.0 | 508.0 | |
| 1000 | ± 8.0 | 992.0 | 1008 | |
After the completion of calibration, calibration label shall be affixed into the respective micropipette. 5.3.9 Record the calibration details in respective format. 5.4 Calculation for calibration Volume calculation Avg. of 10 weighing Mean volume = —————————- x 1000 0.99753 (density of water at 25°c is 0.99753g/ml) The accuracy of the pipette is calculated as followed by, Mean volumes- nominal volumes% Accuracy = ————————————— x 100 Nominal volume The precision of the pipette is calculated as follow by STD deviation % precision= —————————- x 100 Mean volume 5.5 Half yearly by in-house and yearly by outside party. 5.6 10ml micropipette calibration done by outside party in once in year. 5.7 After calibration completion affix a label on micropipette as per calibration respective format. |