My question is related to interpreting results by Flow Cytometry.

Let’s say I have a background level of staining of Isotype-FITC at 100 MFI units. Then, let’s say I have protein A-FITC, which stains at 20,000 MFI units, and Protein B-FITC which stains at 10,000 MFI units. Assume that the staining for A and B are of equivalent affinity and kinetics.

To find the relative levels, do I divide numbers, like this:

A/bkgd = 20,000/100 = 200; B/bkgd = 10,000/100 = 100; A-B = 200-100 = 100 unit difference, and A/B = 2.00 fold difference

Or do I subtract numbers to quantify changes in protein levels, like this:

A- bkgd: 20,000-100 = 19,900; B-bkgd = 10,000-100 = 9,900; A-B = 19,900 - 9,900 = 10,000 unit difference, and A/B = 2.01 fold difference.

Also, is there a reference for this?