Agilent 6530 reference spray and mass accuracy inconsistencies

Hi all,

Our lab recently purchased a refurbished 6530 to go with an Infinity 1290 series LC stack we inherited from the previous inhabitants. Recently, I have been having trouble with maintaining good mass accuracy for certain analytes I am measuring and I believe the problem may lie with the reference spray. Our lab uses two reference ions, purine (121 mz) and HP-0921 (922 m/z). Generally at the start of a run, both reference ions can be found easily. However, as the run progresses, generally only one or none of the ions can ever be found. For example, here is an EIC for purine in the first blank injection of a worklist I had run last evening:

And here is an EIC at the end of the run:

There is a similar story for the second reference. Here are two EICs from the start and end of the worklist:

Similarly, the mass accuracy for the reference ions can be well over 100 ppm in scans where they are not very abundant. Can anyone offer any advice for maintaining a consistent spray for the reference solution? Currently I have tried plumbing the outlet from the pump into the reference sprayer with the valve open to try to flush any sort of clog. Any advice for solving this or maintaining good mass accuracy in general would be greatly appreciated.


  • From the chromatograms it looks like there is some form of clogging, which might either be in the reference sprayer itself, or the tubing itself. You can swap analytical and reference sprayer, and see if the signal gets more stable. If not, then it might indeed be the tubing. Counterflush as you described, or a tubing replacement. In general I recommend adjusting the intensities at 1Hz to be over 100,000 counts at the later part of the LC gradient. With that you should be at worst within 10-20ppm over a 24h temperature cycle. I also recommend changing the acquisition parameter for the reference masses to be within 20ppm

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