ã€purpose】
This regulation stipulates standard operating procedures for the use, calibration and management of pipettes, so that operators and pipette managers can follow the rules to ensure that the use, calibration and management of pipettes meet the requirements of the GLP regulations.
ã€procedure】
1 Use of pipettes
1.1 Select a pipette with a suitable range: the pipette can only accurately take liquid within a specific range. If it exceeds the minimum or maximum range, it will damage the pipette and cause inaccurate measurement;
1.2 Set capacity value
1.2.1 Coarse adjustment: quickly adjust the capacity value to close to your expected value by adjusting the knob;
1.2.2 Fine adjustment, when the volume value is close to the set value, the pipette scale display window should be placed in front of your eyes in parallel, and the volume value can be slowly adjusted to the expected value by adjusting the knob, so as to avoid visual errors Impact;
1.2.3 Precautions when setting the capacity value: When adjusting the range, if you want to change from a large volume to a small volume, you can turn the knob counterclockwise according to the normal adjustment method. However, if you want to adjust from a small volume to a large volume, you should first turn the scale knob clockwise to the scale beyond the range, and then return to the set volume, so as to ensure the highest accuracy of the measurement. In the process of setting the volume value, it is forbidden to turn the button out of the range, otherwise it will jam the internal mechanical device and damage the pipette.
1.3 Assembly of the suction nozzle (pipette tip): Insert the top of the white sleeve into the suction nozzle, while gently pressing down firmly, the pipette in the handle rotates counterclockwise until the suction nozzle is clamped. Remember not to use too much force, let alone use the method of chopping tips to install. The sign that the suction nozzle is tight is slightly beyond the O-ring, and you can see that the connection part forms a clear seal ring. ;
1.4 Pre-washing liquid suction nozzle: after installing a new liquid suction nozzle or increasing the volume value, the liquid to be transferred should be sucked and discharged two to three times to ensure the accuracy and accuracy of the pipetting work;
1.5 Aspiration: Hold the upper part of the pipette with four fingers together, press the button on the top of the stopper with your thumb, press down to the first stop point, and then immerse the tip vertically into the liquid surface for 2 ~ 3mm, slowly and smoothly loosen Open the button, suck up the liquid, and stay for 1 to 2 seconds (longer residence time can be extended for a viscous solution);
1.6 Pipetting: Slowly lift the pipette to take out the suction nozzle to ensure that there is no residual liquid on the outer wall of the suction nozzle. Use qualitative filter paper to wipe out the liquid droplets that may adhere to the outside of the nozzle. Be careful not to touch the mouth of the suction nozzle;
1.7 Visually check whether the volume of liquid inhaled is reasonable;
1.8 Dispensing liquid: stick the suction nozzle to the inner wall of the container and maintain a 20 ° -40 ° tilt, press the button to the first stop point steadily, after stopping for 1-2s (the viscous liquid needs to stay longer), continue pressing At the second stop, the residual liquid is discharged. Release the button, and then move the suction nozzle upwards along the inner wall;
1.9 Press the tip ejector to remove the tip, and the tip must be replaced when drawing different sample liquids;
2 Calibration of the pipette
2.1 In order to ensure the accuracy of sampling, the pipette should be calibrated regularly. The center stipulates that the pipette should be calibrated twice a year, and the second interval should not exceed 7 months, which should be completed by the Logistics Support Department. Pipettes with a maximum volume of less than 10 μL are sent to qualified units for calibration;
2.2 Measurement performance requirements: When the pipette is at a standard temperature of 20 ° C, its volume tolerance and measurement repeatability should meet the requirements of Table 1;
2.3 Calibration conditions
2.3.1 Calibration environment: The pipette should be carried out under the condition that the room temperature is between (20 ± 5) ° C and the change in room temperature is not greater than 1 ° C / h;
2.3.2 Calibration medium: use ultrapure water, put it in the laboratory 24 hours in advance, so that the temperature difference between the temperature and room temperature should not be greater than 2 ℃;
2.3.3 Calibration equipment: The main equipment must be certified by the legal technical institution and within the certification period. Electronic balance (measurement range 200g; division value 0.1mg); temperature and humidity meter; thermometer (measurement range (0 ~ 50) ℃; division value 0.1 ℃); one beaker (50ml); one beaker (100ml); Qualitative filter paper small pieces (after constant temperature treatment);
2.4 Calibration method
2.4.1 Turn on the electronic balance and zero-adjust it at no load; place the thermometer in a vessel containing ultrapure water;
2.4.2 Put in a small beaker and wait for the balance display to stabilize, then press the zero key to return to zero. Install the corresponding pipette nozzle on the pipette, pre-wash the pipette tip, and record the current environmental temperature and humidity on the pipette calibration record table;
2.4.3 Adjust the volume of the pipette to the inspection point;
2.4.4 Hold the pipette vertically and rotate the button to the verification position. At this time, inject the pipette nozzle into the container containing ultrapure water and keep it at the position of 1 ~ 3mm below the liquid surface, slowly release the button and wait Leave the liquid surface after 1s ~ 2s. If the suction nozzle occupies excess liquid, use qualitative filter paper to dry it (be careful not to touch the mouth of the suction nozzle)
2.4.5 Place the liquid outlet of the suction nozzle against the inner wall of the beaker on the balance pan and form an angle of about 45 ° with it. Next, move the suction nozzle upwards along the inner wall of the weighing cup.
2.4.6 After the balance display is stable, read and record the value displayed by the electronic balance on the pipette calibration record table. After recording, reset the electronic balance;
2.4.7 Repeat 5 times to execute (4) ~ (6). After recording, adjust the pipette volume to the next inspection point to continue. The inspection points are adjusted in accordance with the principle from small to large;
2.5 Data processing
2.5.1 Calculation of the actual volume of the pipette: Bring the measured mass value into the following formula to obtain the actual volume value of the pipette at the standard temperature of 20 ℃;
In the formula: V20-the actual capacity of the pipette at a standard temperature of 20 ℃, mL;
m ——Quality of ultrapure water discharged from the pipette being tested, g;
K (t) ——The proportionality coefficient between the measured value m and the ultrapure water temperature during the measurement, listed in Appendix 2;
2.5.2 Calculation of the relative error of the volume of the pipette: Bring the converted actual volume value into the following formula to obtain the relative error of the volume of the pipette at the standard temperature of 20 ℃;
In the formula: V-nominal capacity, μL;
——The arithmetic average of 5 measurements, μL;
2.5.3 Volume repeatability calculation of the pipette: The converted actual volume value is brought into the following formula to obtain the volume repeatability of the pipette at a standard temperature of 20 ° C;
Where: σn-1-standard deviation;
n ——Number of measurements;
S-repeatability;
2.5.3 Fill in the calculation result on the pipette calibration record form and give the calibration conclusion.
2.6 Processing of calibration results
2.6.1 The calibrated pipette is labeled with the calibration certificate, indicating the calibration number, unit, date and validity period. The format of the calibration number consists of ZJ year-pipette number-the number of calibrations in this year. For example, the number of the first calibration of the pipette No. 001 in 2009 is ZJ09-001-01.
2.6.2 If one of the three ranges tested has a capacity tolerance and measurement repeatability that exceeds the requirements of Table 1, it should be cleaned, corrected, and recalibrated. If the re-calibration is still unqualified, it shall be returned to the manufacturer for repair or disposal;
2.7 Correction of the pipette
2.7.1 Adjust the pipette to the minimum range, obtain the current actual volume value according to the calibration method, and adjust according to the difference from the marked volume value through the adjustment knob on the pipette;
2.7.2 When the actual measured volume value and the marked volume value after the adjustment meet the requirements of Table 1, adjust the pipette volume to the middle range and continue the measurement. When the intermediate range meets the requirements, adjust to the maximum range to continue measurement.
2.7.3 If one of the middle range or the maximum range does not meet the requirements of Table 1, it needs to be readjusted to the minimum range to continue the correction. Until the values ​​of the three ranges meet the requirements of Table 1.
2.7.4 The corrected pipette needs to be calibrated again.
2.7.5 If the minimum range is less than 10μL, no correction will be made. If the maximum range and intermediate range meet the calibration requirements, continue to use, otherwise return to the factory for repair.
3 Precautions for the use of pipettes
3.1 If it will not be used for a long time, adjust the range of the pipette to the maximum scale to keep the spring in a relaxed state to protect the spring;
3.2 Regularly clean the outer wall of the pipetting gun, you can use 95% alcohol or 60% isopropyl alcohol, then wipe with distilled water, and dry naturally;
3.3 Check whether there is liquid leakage when using. The method is to suck the liquid and hang it vertically for a few seconds to see if the liquid level drops. If liquid leaks, check whether the suction nozzle matches and the spring piston is normal;
3.4 When there is liquid in the pipette nozzle, it is strictly prohibited to place the pipette horizontally or upside down to prevent the liquid from flowing into the piston chamber and corroding the pipette piston;
Attached Table 1: Tolerance of pipette capacity and measurement repeatability
Nominal capacity / μL
Calibration point / μL
Capacity tolerance
± (%)
Measurement repeatability
≤ (%)
20
2
12.0
6.0
10
8.0
4.0
20
4.0
2.0
50
5
8.0
4.0
25
4.0
2.0
50
3.0
1.5
100
10
8.0
4.0
50
3.0
1.5
100
2.0
1.0
200
20
4.0
2.0
100
2.0
1.0
200
1.5
1.0
1000
100
2.0
1.0
500
1.0
1.0
1000
1.0
0.5
5000
500
1.0
0.5
2500
0.5
0.2
5000
0.6
0.2
Attached table 2: K (t) value
The above are the standard operating procedures for the use, calibration and management of pipettes organized by the editor
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