Testosterone background in UPLC

I am developing an assay for low-level testosterone (low pg/mL range) and am seeing a significant testosterone signal, complete with spot-on retention time and ion transitions,  that we are reasonable certain is coming from the Acquity UPLC. The basal level quants out to 20-40 pg/mL, which is to high for us. We have tried:1. Changing the needle wash solutions2. Changing the needle3. Changing the injection modes4. The signal is present if you do a 0 uL injection at the same levels as a 10 uL injection.5. It is present regardless if you inject solvent straight out of a bottle, or an extracted blank at nearly identical levels.6. Waters has come in and basically rebuilt the entire injection path.7. You can wash the entire system with straight ACN for an extended period of time and then re-equilibrate under initial conditions and get a yet stronger signal than you had before.8.The signal is strongest on the first injection and then diminishes to a 'basal level' by the second and subsequent injections.If anyone has successfully dealt with this problem, please let me know.THanks,
Mary

Answers

  • Thank you for your input. We generally use Optima grade water, but it couldn't hurt to replace everything again. We did have to revert to house DI for a week or so. I'll let you know how it turns out.

    Have a good day!

    M

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