CALIBRATION OF MICROPIPETTE

• PROCEDURE:

• Calibration procedure:
• Operate the Micropipette as per SOP. The micropipette shall be calibrated based on dispensing range.
• For 20 µL -200 µL

Set volume             Permitted Volume

20 µL                         19.8 – 20.2 µL

100 µL                        99.2 – 100.8 µL

200 µL                        198.8 – 201.2 µL

Enter Details in Calibration record as per Format-I.

• For 100 µL -1000 µL

Set volume in                   Permitted Volume

100 µL                                99.2 – 100.8 µL

500 µL                                 496 – 504 µL

1000µL                                994 – 1006µL

Enter Details in Calibration record as per Format-II.

• For 500 µL -5000 µL

Set volume in             Permitted Volume

500 µL                         496 – 504 µL

2500 µL                       2484 – 2516 µL

5000µL                         4970 – 5030µL

Enter Details in Calibration record as per Format-III.

• For 1000 µL -10000 µL

Set volume in                   Permitted Volume

1000 µL                            992 – 1008 µL

5000 µL                            4960 – 5040 µL

10000 µL                          9940 – 10060 µL

Enter Details in Calibration record as per Format-IV.

• The water which is to be aspirated is taken into a suitable vessel.
• Attach suitable tips to the pipette firmly.
• Press the control button to the first stop, immerse the pipette tip into the water.
• Allow the control button to slide back slowly.
• Dispense the aspirated water into the weighed bottle.
• Similarly, do the operation for five times and collect the water into the weighed bottle.

• Divide the average weight of water with weight/mL of water at 25°C and compare it with the permitted volume.
• Calculation of weight/mL of water:
• Take a 25 mL pycnometer and note down the empty weight(W1)
• Fill the pycnometer with water, close with stopper provided and wipe off the excess water.
• Now note down the weight of pycnometer with water at 25°C (W2) Weight/mL of water = (W2-W1) / 25.
• Calibration shall be carried out preferably at temperature of water at 25°C. If temperature varies, observed temperature shall be noted and the respective density of water shall be considered for calculation as per below table instead of calculating weight/mL of water.
• Enter Details in water calculation record as per Format-V.
• The fixed volume micropipette shall be calibrated as per Format-VI.

Table-1
Density of Water (g/mL) vs. Temperature (°C)

Whole degrees are listed down the lefts hand side of the table, while tenths of a degree are listed across the top

Tem per atu re	0.0	0.1	0.2	0.3	0.4	0.5	0.6	0.7	0.8	0.9
	Density of water
15	0.9991	0.999084	0.99907	0.99905	0.99904	0.99902	0.99901	0.99899	0.99898	0.99896
16	0.99894	0.998926	0.99891	0.99889	0.99888	0.99886	0.99884	0.99883	0.99881	0.99879
17	0.99877	0.998757	0.99874	0.99872	0.9987	0.99869	0.99867	0.99865	0.99863	0.99861
18	0.9986	0.998576	0.99856	0.99854	0.99852	0.9985	0.99848	0.99846	0.99844	0.99842
19	0.99841	0.998385	0.99837	0.99835	0.99833	0.99831	0.99829	0.99827	0.99824	0.99822
20	0.9982	0.998183	0.99816	0.99814	0.99812	0.9981	0.99808	0.99806	0.99804	0.99801
21	0.99799	0.99797	0.99795	0.99793	0.9979	0.99788	0.99786	0.99784	0.99782	0.99779
22	0.99777	0.997747	0.99772	0.9977	0.99768	0.99766	0.99763	0.99761	0.99759	0.99756
23	0.99754	0.997514	0.99749	0.99747	0.99744	0.99742	0.99739	0.99737	0.99735	0.99732
24	0.9973	0.997271	0.99725	0.99722	0.9972	0.99717	0.99715	0.99712	0.9971	0.99707
25	0.99704	0.997018	0.99699	0.99697	0.99694	0.99691	0.99689	0.99686	0.99684	0.99681
26	0.99678	0.996756	0.99673	0.9967	0.99668	0.99665	0.99662	0.99659	0.99657	0.99654
27	0.99651

Example: To find the density of water at 16.1 °C, you would first find the whole degree by searching down the left hand column until you reach ’16’. Then you would slide across that row until you reach the column labelled ‘0.1’. The density of water at 16.1°C is 0.998926 g/mL.

• ANNEXURES: 

ANNEXURE NO.	TITLE OF ANNEXURE
Annexure-I	Calibration record for 20 – 200 µL micropipette
Annexure-II	Calibration record for 100 – 1000 µL micropipette
Annexure-III	Calibration record for 500 – 5000 µL micropipette
Annexure-IV	Calibration record for 1000 – 10000 µL micropipette
Annexure-V	Weight/ mL of water calculation record
Annexure-VI	Calibration record for fixed volume micropipette

Annexure-I
CALIBRATION RECORD FOR 20 – 200 µL MICROPIPETTE

Annexure-II
CALIBRATION RECORD FOR 100 – 1000 µL MICROPIPETTE

Annexure-III
CALIBRATION RECORD FOR 500 – 5000 µL MICROPIPETTE

Annexure-IV
CALIBRATION RECORD FOR 1000 – 10000 µL MICROPIPETTE

Annexure-V
WEIGHT/ ML OF WATER CALCULATION RECORD

Annexure-VI
CALIBRATION RECORD FOR FIXED VOLUME MICROPIPETTE