Mascot, Biotools and Flexanalysis ms/ms

[chemstation]

Hello,
I'm trying to find out from someone who uses or knows,
how Flexanalysis, Biotools and Mascot work together in ms/ms (particularly in batch mode)
the question relates to how and when flexanalysis uses the information it gets back from the mascot server. (we have a Bruker MALDI with flexanalysis & Biotools but no in-house mascot server.)

eg if we create a sequence that has an ms and a ms/ms method in each line,
does flexanalysis do a ms/ms immediately after each ms acquisition ?, or
does it wait until the end of the sequence, complies the results from mascot into one big file, and then does a ms/ms based on the result of the one big file???

thanking you in advance

Alex

[Kostas Petritis]

Alex,

There is not a predetermined way to use the software, it depends on what you want to do and what do you know about the biology.

For example, in biomarker discovery, if you want to find lets say cancer markers through proteomics or metabolomics, you want to search for metabolites or peptides/proteins that change in abundance from one state to another. Due to undersampling issues, and problems with compound identifications (i.e. especially in the case of metabolites or peptides/proteins with post-tranlational modifications) it is impractical to do MS/MS of all peptides/proteins/metabolites. Instead, you can do LC-MS by using the accurate mass information of the cancer and control sample, then use flexanalysis/biotools to compare the same "features" and identify the ones that significally change from one state to another. Once you know the "feautures" (i.e. mass, time, isotopic distribution and/or fraction number) that have shown an expression difference, you can compile and inclusion list of the feauture that you are only interested and then require LC-MS-MS analysis of the subset of compounds that you are interested.

You can then use Mascot and Biotools in order to try to identify peptides/proteins/metabolites that showed expression differences and these will be your potential biomarkers that you will have to further validate by other means (that is another story all together).

A case study with the tools that you mentioned can be found at Cheng et al. Clinical Chemistry 51:12 2236-2244.

Having said all these, I have not personally used Flexanalysis or Biotools as we do not have Brucker ToF's in house but we have similar informatics in-house tools to do that.

As of now, there is nothing that you can use to do this on the fly (i.e. differencial expression, identification of compound of interest and targeted fragmentation of the feature of interest). Propably you will see proteomic/metabolomic platforms like this in the not too distant future (i.e. < 10 years)... the exact way to do it is still to be determined (there are several possibilities)...