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No fragmentation in ESI

Posted: Tue Feb 19, 2019 6:16 pm
by MrGC
Dear all,

As you can gues from my title I do not experience any fragmentation in MRM mode while tuning.
My normal tuning for an appropriate MRM method looks as follows (maybe there are already a few flaws in there):
1) I directly infuse the analyte of interest by a syringe.
2) tune the machine i.e. I regulate cone voltage, capillary voltage, LM, HM and so on
in between I do several MS scans to see if I achieve improvements regarding sensitivity
3) If my sensitivity is good enough I switch on the collision gas which gives a hissing sound. The signal of my precursor ion gets lower depending on the intensity I set for the collision but I cannot see any fragments.

We have the same machine a second time (waters quattro micro API) and my approach works on it i.e. I do see fragmentation of the exact same saple.
Do you have any Ideas? Or some advice?

Re: No fragmentation in ESI

Posted: Fri Feb 22, 2019 4:45 am
by GoinBoardin
Is this a triple quad instrument, with a q2 being a collision cell? Sorry, I am not familiar with the instrument you have.

What is your collision gas pressure? Is this hissing normal for this instrument (does the other one you have do it too?)? My old Thermo TSQ7000 doesn't make a sound other than a solenoid valve opening, there is no hiss. A hiss sounds like a leak to me. Can you monitor the pressure in the CID cell, with collision gas on vs off?

My best guess is a leak of collision gas causing insufficient collision gas pressure in the collision cell and very low fragmentation efficiency.

My other thought is on the ability of the final quad to transmit the ions of interest. When was the last calibration? Could the system be dirty? In full scan mode, is there much difference when using Q1 to scan vs Q3 to scan? That is, if that is possible on this instrument. Rather than an MRM, what happens with a Product ion scan?

Best of luck in your troubleshooting.

Re: No fragmentation in ESI

Posted: Sat Feb 23, 2019 7:07 pm
by Ian_R
MrGC wrote:
Dear all, I switch on the collision gas which gives a hissing sound. The signal of my precursor ion gets lower depending on the intensity I set for the collision but I cannot see any fragments.


It would probably do that if the internal gas line to the collision cell is broken/disconnected. Adding gas will just cause poor analyzer vacuum and low sensitivity rather than fragmentation inside the collision cell. Is the analyzer vacuum normal?

Re: No fragmentation in ESI

Posted: Fri Mar 01, 2019 7:51 am
by mckrause
You should not hear anything when you turn on the collision gas. If you have a hissing sound then you most likely have too much collision gas pressure at the back of the instrument. What do you have your tank pressure set to? Note that the recommended pressure is around 7-10 psi (yeah, really, really low).

It is also possible that you have a broken line to Q2, although if that were the case I would think that you would see the issue in your tuning and optimization.