all of a sudden, no elution
Posted: Sat Mar 28, 2015 3:35 am
Hi, guys
I am a newbie to UPLC/HPLC (<3 wk experience). Starting to run a new assay to quantitate a compound using fluorescence detector (exc: 370nm, em: 460nm). Using 80:20 (0.2% formic acid, 100% methanol) solvent system.
My compound used to elute at 6 min run (for week 1). All of a sudden, it stopped coming out throughout the 12 min run cycle during the past 4 days.
I thought I screwed up the system, but when I ran a standard compound (exc: 405nm, em:619nm), the standard ran fine (under 92:8 (ammonium acetate, acetonitrile)) so I assumed the fluorescence detector/lamp is fine.
I might have prepared a 2% formic acid, instead of the 0.2% formic acid solvent. Is it possible that 2% formic acid could screw up my elution? If not, what could other possibilities be that all of a sudden I am getting no elution.
terrified as the samples are precious and I could not keep testing the system using the samples. Any suggestion will be greatly appreciated. Thank you very much.
I am a newbie to UPLC/HPLC (<3 wk experience). Starting to run a new assay to quantitate a compound using fluorescence detector (exc: 370nm, em: 460nm). Using 80:20 (0.2% formic acid, 100% methanol) solvent system.
My compound used to elute at 6 min run (for week 1). All of a sudden, it stopped coming out throughout the 12 min run cycle during the past 4 days.
I thought I screwed up the system, but when I ran a standard compound (exc: 405nm, em:619nm), the standard ran fine (under 92:8 (ammonium acetate, acetonitrile)) so I assumed the fluorescence detector/lamp is fine.
I might have prepared a 2% formic acid, instead of the 0.2% formic acid solvent. Is it possible that 2% formic acid could screw up my elution? If not, what could other possibilities be that all of a sudden I am getting no elution.
terrified as the samples are precious and I could not keep testing the system using the samples. Any suggestion will be greatly appreciated. Thank you very much.