Hi everyone,
I am working on a protein in macrophage cells. My aim is to discover its interacting partners through LC-MS/MS. I have two conditions. One is where the protein stays intact and pulled-down with its interactors (condition A), and the other condition yields cleaved protein hence no/less interacting partner (Condition B).
I run my samples in LC-MS/MS and then analyse it with MaxQuant. The question I have is which parameter to use after MQ analysis. Ideally I’d like to see an enrichment of the interacting protein (also my immunoprecipitated protein) in my condition A, compared to condition B. I am not sure whether I should be using iBAQ or LFQ values to further analyse the samples in Perseus. I’d be happy if anyone can share their ideas.
Thanks
In IP-MS experiments you normally have quite a lot of unspecific background binding, that is sufficient for LFQ normalisation, so I rather use LFQ then ibaq. In both cases, you need to address the question of proteins, that are not quantified in mock/ condition B. You can do this either by follow-up data-analysis with specific tools for IP-MS or if you use Perseus, you can imputate values for condition B and do a one sided T-Test, looking only for enrichment in A.
Dear Florian Bonn
Thank you very much for your answer. I use Perseus and will try one-sided ttest tomorrow as you suggested.
Do you have a suggestion of any software specifically useful for IP-MS ?
Thanks!
Arda
No, I do not have personal recommendations, but saw recently some papers, but I do not have anything specific in mind.
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