HI everybody,
I'm performing qPCR data analysis and a colleague suggested me a sort of modification over 2^-ddCt calculation. I have the average dCt of four experiment, control and treatment. He suggested instead of using the control value in each experiment, to use in all the experiments the average of controls. Doing this I obtain controls around 1 and not exactly 1. Does it make sense to you? Is this a validated way to make calculations in order to reduce inter experimental variability?
Thanks for your answers
Laurence Stuart Dawkins-Hall
- No !!
- Do not average across plates
- Your calibrator or control sample should ALWAYS be '1'
- You can look at interplate variability but you must take the data initially plate by plate
- The only way you can accurately correct for inter plate variability it to use a plate (extra) control on each plate which can then be used to adjust for plate to plate variation so that data can be pooled and meta analysis can be performed
- Here is a a link to pertinent info on how to analyse multiple plate data by meta analysis:
- https://www.researchgate.net/post/qPCR_analyses_involving_in-between_plates
Article Removal of between-run variation in a multi-plate qPCR experiment
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