how to choose SRM specific transitions ?

[Cassandre-2015]

Dear all,
I have a general question.
I am using gcmsms but I guess it can be the same for lcmsms.
how do you select the precursors and daughter ions of the srm transitions you want to develop ?

I understand it has to be as specific as possible.
But if there are several possible precursors and then several daughter for each, how do you choose ?
It can be time consuming and I would be very interested to know your hints and tips about that .

many thanks

best regards

[James_Ball]

LCMS is a little easier because you can infuse a solution of the target analyte while finding and optimizing the parameters.

For LCMS I begin with a full scan on MS1 and look for the molecular ion, or an adduct with good sensitivity. Then I fix MS1 on that mass +/-3amu and optimize the sensitivity. Then I run MS1 locked on that mass with collision cell ramping through a good range of energies and scanning MS3 and seeing what daughter masses are created. Select a few strong daughter masses then run all the cell parameters through their ramps while locked on the parent with MS1 and locked on the daughters with MS3 and finding the optimum settings for each parameter for each daughter, then you are finished.

For GCMS I would think it is similar except you have to do multiple injections instead of infusing the analyte. Most good MS software with somewhat automate the task, but that is what it is doing in the background.

[lmh]

the only thing that I'd add to James' reply, is that when you've got a short-list of useful transitions, it's worth running some real samples while monitoring all the transitions (more transitions than you intend to use, ultimately). You will probably want several usable transitions, so you have a quantifier and one or two qualifiers, depending how you work. It is depressing how often, in real samples, a beautifully promising transition gives an array of peaks that makes a cross-section of the alps look flat. You may sometimes find that you have to reject one of your best initial transitions because it lacks specificity in real samples, and fall back on a less abundant transition that gives a clear peak well separated from any neighbours.

[mckrause]

Couldn't agree with lmh more. Nothing more disappointing than having a promising daughter ion completely wiped out by an interference from your sample.

(Still laughing about the "cross-section of the Alps look flat"). OK, now I wiped the tears of laughter away. If you have a relatively constant matrix, such as with quality control on a product, then you only have to run the experiment one time to set your parameters. If you have a highly variable matrix like food, then you really have to be careful and spike all samples. I'll leave the theoretical explanation to better minds than mine, but operationally you will see quite a large variation in daughter ion production depending upon your matrix, and thus variations in your recovery. We spike each and every sample when we run either LC-MS/MS or GC-MS/MS with all analytes of interest, not just internal and surrogate standards.