What value should I choose for the downstream analysis in proteomics?

Question

[SayHey]

Hi there,

I am a very beginner to proteomics. Recently, I obtained a bunch of proteomics data. But I have no idea of how to analyse it. I have read some tutorials. They performed the analysis based on spectral count or intensity. I could not find it in my dataset. So my first question is what value should be used. If the value has been chosen, what is the proper method for the data?

My dataset looks as follow:

Many thanks Say Hey

Answer 1

[Michael]

This looks like an output from Proteome Discoverer by ThermoFisher the manufacturer of the MS instrument used for the analysis. The analysis is already done and proteins have been identified, like Keratin. So, now you know Keratin was really there in your sample and it has about 62% coverage by peptides, identified by the Sequest engine. If you want to do another analysis using a different tool you should ask for the data in .raw format.

[SayHey]

Thank you for your answer, Micheal.

I have another question about it. If I have two or more tables like it from different samples, is it possible to do a differential analysis? When I have data in .raw format, what should I do for other analysis? Such as differential expression analysis, protein-protein interaction.

[Michael]

I am not so much an expert in proteomics and I hope others will correct me if I am wrong. What I would say to you is that the experiment and analysis need to follow the biological question you are trying to answer. Therefore, I cannot tell you what you should do. On the other hand, what you can further do with the data may be quite limited by the technology used. You seem to look for a quantitative approach, but peptide mass fingerprinting is not really that. It's good at doing one thing, identifying proteins, but how much protein there is in a complex mixture is much hard to conclude from such a simple setup because there are so many factors that play into peak intensities and general detection.

I think you should talk to your facility manager about the experimental design and their capabilities to perform real quantitative proteomics experiments and subsequent analysis steps. If you find a good service provider, please let me know. On a general basis, it is strongly advised to formulate the question before doing the experiments.

[SayHey]

Thank you for your advice.