TMT with fractionation

[SheonMary]

Hi,
I am new to FragPipe and want to clarify if I am using the right workflow and parameter. Experimental design consists of TMT-10plex samples combined and then offline fractionated into 9 fractions. However, there are two such combinations each with 9 fractions (see the table below). In FragPipe, I am using TMT-10 workflow. Do I need to change any other parameter so there is at the end two comparision 1. comparison between 2 groups (HM vs C) and 2. comparison between 4 groups (mHM, mC, fHM, fC).

[fcyu]

You need to

1. Assign the first 9 fractions to one "experiment", and assign the second 9 fractions to another "experiment".
2. Edit the annotation files in the "Quant (Isobaric)" tab to annotate the TMT channels of two "experiments" according to the names showing in your table.

Since your sample doesn't have reference sample, you could select the virtual reference option in the "Quant (Isobaric)" tab.

Here is a tutorial (https://fragpipe.nesvilab.org/docs/tutorial_tmt-2plexes.html) in case you need it.

Best,

Fengchao

[SheonMary]

Thank you so much.