Work in groups of three. One person in the group should work with the P-20, another with the P-200 and the third with the P-1000. Each person should prepare three identical samples and then determine the A590 of the three samples. From the data, you will be able to determine if the micropipette is measuring volumes correctly.
1. Each person in your group of three will work with a different micropipette and perform the same transfers in triplicate, as detailed below. The final volume (water + BPB) in each tube will be 1.0 mL. Calculate the volume of water that will need to be combined with each of the following to give 1.0 mL, and record your calculations in your lab notebook:
Group member A: Use the P-20 to transfer 10 μL of 0.05% BPB to ____ μL of water. Group member B: Use the P-200 to transfer 100 μL of 0.005% BPB to ____ μL of water. Group member C: Use the P-1000 to transfer 300 μL of 0.005% BPB to ____ μL of water.
2. To minimize our plastic waste, strategize how to minimize the number of tips that you use without contaminating the stock solutions. A tip can be used multiple times, but a tip that has been used for BPB cannot be used to subsequently transfer deionized water. Combine the BPB solution and water to give a final volume of 1.0 mL.
3. Measure the A590 of the three solutions with the spectrophotometer and record the data in your notebook.
- Compute the mean and standard deviations for your three measurements, using either a calculator or Excel. The standard deviation reflects the precision of your measurements.
- Enter these values on the chart that your TA has prepared on the whiteboard. Compare the values that your group obtained with each of the three pipettes with the aggregated class measurements. If the averages that your group obtained are significantly different than those that other groups obtained with the same micropipette, your micropipette may not be transferring volumes accurately.
Notify your TA if any of the micropipettes are not performing properly. Your TA will follow up on your observations and test the micropipettes with the gravimetric test described earlier in the chapter.