Yaamini’s Notebook: SRM Dilution Calculations

My prior calculations were incorrect!

As I mentioned previously, Laura and I need to run dilutions of our samples to confirm that as the concentration of protein in our samples decreases, the assay does not detect them as intensely. When I showed Emma my calculations, she suggested making dilutions based on protein concentrations (µg) as opposed to µL of sample added. Here are my final numbers:

Table 1. Dilution calculations

Vial Dilution Ratio µg to Inject µL to Inject µg oyster Needed µg geoduck Needed µL oyster needed ((µg/µL)x15 µL/1 µg/µL) µL geoduck needed ((µg/µL)x15 µL/2 µg/µL) PRTC added µL ACN Total Volume (µL)
1 10:1 1 1 0.9 0.1 13.5 0.75 1.5 N/A 15.75
2 7.5:1 1 2 0.87 0.13 6.5 0.5 0.75 7.25 15
3 5:1 1 2 0.8 0.2 6 0.75 0.75 7.5 15
4 2.5:1 1 2 0.6 0.4 4.5 1.5 0.75 8.25 15
5 1:2.5 1 2 0.4 0.6 3 2.25 0.75 9 15
6 1:5 1 2 0.2 0.8 1.5 3 0.75 9.75 15
7 1:7.5 1 2 0.13 0.87 1 3.25 0.75 10 15
8 1:10 1 1 0.1 0.9 1.5 6.75 1.5 5.25 15

For PRTC, our stock concentration volume is 0.5 pmol/µL. I changed the injection volume for the 10:1 and 1:10 dilutions from 2 µL to 1 µL to ensure we were pipetting volumes over 0.5 µL. This made it so that the final volume for the 10:1 injection was 15.75 µL, but Emma said that was okay.

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