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Background variation


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4 replies to this topic

#1 Gradstudent78

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Posted 31 March 2011 - 01:30 AM

I ran a bunch of blank samples in a plate and noticed that towards the end of the plate samples are having a higher background then samples at the beginning of the plate. For example the blanks in wells A1 and A2 average ~0.058 optical density while H9 amd H10 ~0.083. What could be responsible for this? Thanks.

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#2 PAO_ahac

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Posted 31 March 2011 - 04:27 AM

What is your %CV of replicates? Could be assay variability, carry over, manual v. automated washing, insufficient washing etc.

#3 Gradstudent78

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Posted 31 March 2011 - 05:44 AM

What is your %CV of replicates? Could be assay variability, carry over, manual v. automated washing, insufficient washing etc.


%CV for replicates are all < 10%. I use an automated 4x washing.

#4 mdfenko

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Posted 31 March 2011 - 08:27 AM

does your reader read the whole plate at once or does it read a row at a time?

do you kill the color reaction or just read it?

if the reader does a row at a time and you don't kill the reactions (all at the same time) then the reaction continues during reading.
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#5 Gradstudent78

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Posted 31 March 2011 - 09:30 AM

does your reader read the whole plate at once or does it read a row at a time?

do you kill the color reaction or just read it?

if the reader does a row at a time and you don't kill the reactions (all at the same time) then the reaction continues during reading.


Not sure about the reader (it's fairly old, probably going to buy a new one soon), but we do use a stop solution to kill the reaction.

Edited by Gradstudent78, 31 March 2011 - 09:37 AM.





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