Fluorescence Detector (Waters 2475) Issues
Posted: Mon Apr 10, 2023 7:42 pm
Hello All,
I have a lot of chromatography experience with PDAs, but none with Fluorescence detection. We are analyzing BaP with a method that has been used for years. This is the most bizarre thing I have ever seen. Using an Agilent Zorbax Eclipse XDB-C18, 4.6 x 50mm, 1.8um column. Isocratic Elution, 7.5 min run time, 10uL injection, our area counts just continually climb. We start around 6 million and end up at 8 million. Same exact vial is being pulled from for these injections. So, the %RSDs for the area make the run unusable. If we start a second run, piggyback on the first one (say it starts injecting 9 at night while we are all gone) the area counts are finally stable enough for that run to pass. A few days later, we start the torture all over. First run is trash, but the second one is good. We've ruled out carry over. We've had a technician out to look at it. He says there is nothing wrong with it. His theory is column active sites, but that doesn't make any sense to me either, we are running on the exact same column. 100s of injections. We've looked at a second column. We get the same results. ANY and ALL help would be greatly appreciated.
Jen
I have a lot of chromatography experience with PDAs, but none with Fluorescence detection. We are analyzing BaP with a method that has been used for years. This is the most bizarre thing I have ever seen. Using an Agilent Zorbax Eclipse XDB-C18, 4.6 x 50mm, 1.8um column. Isocratic Elution, 7.5 min run time, 10uL injection, our area counts just continually climb. We start around 6 million and end up at 8 million. Same exact vial is being pulled from for these injections. So, the %RSDs for the area make the run unusable. If we start a second run, piggyback on the first one (say it starts injecting 9 at night while we are all gone) the area counts are finally stable enough for that run to pass. A few days later, we start the torture all over. First run is trash, but the second one is good. We've ruled out carry over. We've had a technician out to look at it. He says there is nothing wrong with it. His theory is column active sites, but that doesn't make any sense to me either, we are running on the exact same column. 100s of injections. We've looked at a second column. We get the same results. ANY and ALL help would be greatly appreciated.
Jen