Hi,
Could anybody please help with the basic understanding how to process AP-MS (affinity purification mass-spectrometry, e.g. mass-spec done on the pull down with an antibody of interest) data. I have 6 datasets: 3 "experiments" and 3 controls (IgG pull downs). Each datafile contains names of the proteins (and gene ID), number of unique peptides, significance level etc. Some proteins were found 1, 2 or 3 times in the experimental samples and other proteins 1, 2 or 3 times in the control samples. Some proteins were found in experimental and control samples (contaminants). I can not understand if I shall just discard contaminants and submit the combined list of all identified proteins to some sort of pathway analysis software or I have to use a statistical package to make a ranged (sorted by a score) list of the identified proteins.
Thank you in advance for all your comments!
Hi,
You can analyse your AP_MS Data using TPP-Trans Proteomic Pipeline for proessing and identification.The sequence can be checked in "RefSeq Database".
There is also an open source software called "SAINT" at http://saint-apms.sourceforge.net/. This will process both spectral count and intensity based data.This uses immunoprecipitation data-purification without unrelated proteins.
You can try with a software called MaxQuant which is a proteomic software package helps out with large data sets analysis from mass-spectroscopy.It also accompanies Perseus for downstream data analysis.
Among all ProHits is a better option as it has data management including storage,MS analytical tools,Visualization of protein-protein interaction.It is also integrated with SAINT.
All the best.
Hi,
thank you very much for your answer. It looks like SAINT is a good choice for my task. I only missing spectral count data to create input files. I have mzdi files as the mass spec result file (and exce files). I did not see where I can use intensity based data In Saint. Do you know how I can extract spectral count from *.mzdi file (I read that mzdi does not have spectral count by itself. I have supporting files to extract that data from).
Hi
Can you tell me the exact file extension? This seems to be different.
Files having extension of .mdi can be read statistically using QSPEC in which variety of extensions can be analysed and protein properties are also framed.
Yet another method would be like if you have a raw data from mass spectroscopy ,then that can be converted to a .dta Files using ExtractMSn Version 3.
All the best
Aishwarya Amirdharaj Hi, thank you very much for you reply.
My files are *.mgf and *.mzid. I also have excel files - report files produced by PEAKS studio. I figured out where my spectral counts are. I prepared and loaded a file into Crapome and it did some analysis. Although I have to learn yet what the output means.
What I see now that I shall not exclude proteins from the binding partners list even if they were picked up in controls (although in the very low quantity - lower spectral counts in fact). I wanted to ask you, so that means we can not exclude proteins pulled down in IgG IPs. We only can compare how abundant proteins are in control and test samples and then make a conclusion. IS that correct?
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