Signal drops during runs

Hello all,

We have having problems with our LC/MS with the TIC signal dropping below the baseline at various points during many runs (see image), including in "blank" samples/washes (both positive and negative mode).

Our setup: 1290 Infinity II LC, diamond hydride silica HPLC column (solvents H2O and acetonitrile, 0.2% acetic acid), with 6545 QTOF (detecting m/z range 50-1200).

We have tried a new column, different batches of solvent, and re-tuning but no luck...


(Please excuse my inexperience, I am new to this world).



  • Hi

    First check purity of nitrogen.if it's contaminated then signal can drop.

    Second thing temperature variation of ionization source.

    If still problem then better to call your local agilent support to troubleshoot onsite.

  • Makes sure that the spray for the reference nebulizer is not sputtering when introducing reference stock. 

  • .I would also check the spray from the main nebulizer, to see if this is sputtering too.  Also, check the wide exhaust tube from the spray chamber and the waste bottle that it is connected to.  Is the flow obstructed by liquid or some other blockage?


  • Apologies for potentially redirecting a discussion, but we have a somewhat similar signal loss challenge with our 6545 QTOF.

    We express recombinant proteins in E Coli, which take the form of inclusion pellets which in turn gets solubilized in guanidine before loading onto a RP column for LC-ESI-MS.

    All is well with most recombinant proteins, except one: there is just one protein that is guaranteed to compromise our performance two ways:

    1) the signal count for our CalMix B (prepared from ESI-L with dilution per maintenance guide with acetonitrile/ HP-0321/water) would drop by >75%

    2) We'd notice mass adducts of ~25Da that trail our CalMix peaks as called out with the red arrows

    We'd regain our performance after an FSE stop by to clean the quad, since cleaning the source or cleaning (and even replacing) the glass capillary does not give our signal back.

    We've pretty much ruled out dirty LC lines and bad CalMix (by flushing LC lines with IPA and water, and prepping/ordering new CalMix); besides, we can carry out LC-MS runs with any other samples----until we get a particular type of recombinant protein.

    The lines of evidence point to the fact we're suffering from something in the sample background matrix, that manages to fly right through the glass capillary and compromises our system (S/N reduction plus mass adducts).

    While we are investigating ways to clean up the sample matrix offline, we're currently dependent on getting an FSE to clean every time we run into the problem.  That is not a sustainable solution, not to mention I'm embarrassed about how dirty our samples are.

    I've been told the "deepest" part an operator should clean is the capillary.  Does the forum know of any other ways to clean any part inside?

    I'm new to Agilent QTOFs; with Thermos, I would have performed a bake-out.  Are there similar functions for Agilent QTOFs?  Thanks for your advice.

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