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need help to resolve the issue on diluted samples

Posted: Wed Oct 18, 2017 4:41 pm
by Ann2017
during the assay validtion we performed the dilution integrity test (5x, and 10x) and the results were good. When we ran the clinical samples which are above the ULOQ, we diluted the samples with 5x in one day and 10 x in another day. (diluted using human blank plasma, same matrix). Each day we have calibration standards, QC low (4 replicate) QC medium (4 replicate);, QC high (4 replicate), and dilution QCs (4 replicate) before we ran our clinical samples. All QCs, including dilution QCs, were passed, but we found that for each sample, the concentration without any dilution < concentration diluted 5 x < concentration diluted 10x.
For example, the measured concentration of one sample is 2830 ng/ml; it changed to 3570 ng/ml using 5x dilution; and 4550 ng/ml using 10x dilution. i don't know why it keeps increasing after it was diluted and back calculated. please advise how to resolve this issue. Thanks!

Re: need help to resolve the issue on diluted samples

Posted: Wed Oct 18, 2017 5:38 pm
by Rndirk
Which detection?

How do you quantitate? Calibration curve? Internal standards? ..

Re: need help to resolve the issue on diluted samples

Posted: Wed Oct 18, 2017 5:58 pm
by Peter Apps
What is your upper quantitation limit ? Were the samples above it ? - if so the measurement result you have for the samples is wrong (and probably low if the calibration curve flattens at the top). If the samples were not above the UQOL, why did you dilute them ?

Peter