Here is my ELISA assay.
Below the picture is the data.
Question:
1) The dilutions from dilution factor (DF) 20, 40, 80 can be converted to a similar values of non-diluted concentrations (38, 39, 37), but the % spike recovery of DF 20 is only 42%.
2) The % spike recovery of DF 160 is 95%, but the converted non-diluted concentration is 23, significantly different from others.
3) Mean 1 = (38+39+37)/3
Mean 2 = (39+37+23)/3
Which mean should I report? Thanks.
Undiluted samples gives less recovery due to hook effect. Samples having higher concentration may create decrease in signal. And the assay type and assay ranges should be establish properly as per the instrument threshold.
If the concentration of analyte in samples are too high than the instrument threshold can determine, then it can create an interference in exact recovery.
It's recommended to first establish the dilution linearity with known concentration before analysing the unknown samples.
Hope it helps to find the answer.
The undiluted samples are higher than your range obtained by the calibration curve. Therefore I suggest to prepare a matching calibration curve.
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