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negative control readings

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#1 Angeline



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Posted 18 June 2018 - 08:34 PM

Hi. I would like to look into the effect of a compound on treated and untreated cells. For my negative controls (comprising of untreated cells), upon staining with antibody, I get a range of 1 to 2% antigen positive cells. My doubt is, whether such a 1 to 2% reading normal as my conceptual is that negative controls should not have any reading as it did not contain the expression. Thank you.

#2 rkay447



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Posted 22 June 2018 - 09:30 AM

It honestly depends on your target and your experimental conditions.  Negative controls are not always completely negative in signal but rather should represent "background" in a system that is known to either be completely missing or significantly lower than your experimental conditions.  For example, a negative control for an antibody can be siRNA mediated knockdown of the target.  You are not going to completely lose all signal as knockdown is not 100% but there should be enough of a reduction in signal to be able to identify a difference in the control non-treated.  If you know for certain that the target is completely absent, then what you are getting is background and can be used to set a limit or gate your analysis as negative results for all other samples. 

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