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Is there a maximum blocking time?


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2 replies to this topic

#1 2xzwei

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Posted 26 April 2013 - 01:52 AM

Hi!
I prepared a 96 well plate with capture AB and over night incubation @ 4°C.
I washed twice and put PBS+10%FCS in each well to block. Unfortunately I cannot continue work today, so is it possible to leave the blocking solution on the wells @ 4°C for 2 days or might this be detrimental? As I already incubated with capture AB I don't want to discard the plate...
What can I do?

#2 swebus

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Posted 26 April 2013 - 02:00 AM

I would not recommend to do this.

I have tried it before - however longer, for about 3 days - and have gotten a lot of background and artificial signaling.

If you use this plate, make sure that you have a negative control and a blank well.

/Peter

#3 Missle

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Posted 26 April 2013 - 05:33 AM

It is not ideal but I've done it. If you have the appropriate negative controls I wouldn't worry too much about background - at least that hasn't been what I've seen. In my experience, you're more likely to see a decrease in signal compared to a more standard incubation time. I think this has to do with loss of capture Ab from the plate.
I guess the real question is how precious are the rest of the samples/reagents for the rest of your assay? If you can't risk repeating the entire assay then discard them now......
good luck!




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