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If Immune sera is showing same reaction to antigen as for plate without antigen,


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6 replies to this topic

#1 Inbox

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Posted 05 July 2013 - 11:16 AM

If Immune sera is showing same reaction to antigen as for plate without antigen, what could be problem?
Thanks.

#2 mdfenko

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Posted 05 July 2013 - 03:01 PM

poor blocking?
talent does what it can
genius does what it must
i do what i get paid to do

#3 Inbox

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Posted 05 July 2013 - 08:23 PM

But Negative sera does not show same reaction to antigen as for plate without antigen. Negative sera behaves negative, positive sera like positive on plate without Ag.

Tried Blocking with Skim Milk powder, Casein. 100 ul blocking solution.

#4 Missle

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Posted 08 July 2013 - 05:51 AM

It sounds as if the immune serum is cross reacting with the block. What is the immunogen?

#5 Inbox

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Posted 08 July 2013 - 11:40 AM

immunogen is Viral protein. I suspected that antibody surface may be matching with Plate surface, but its too much vague.

#6 Missle

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Posted 08 July 2013 - 12:35 PM

If your plate surface is well blocked then the antibody is reacting with the block - not the plate. It would be unlikely that the antibody would be reacting specifically with the plate and any non-specific reaction should be happening with your negative sera too.
I'm not sure what is happening but I would try a couple different blocks (BSA or Fish Serum?). I've worked with antibodies that give lots of problems with a particular block so by default if we get your results we try several different blocks...

Good Luck!!!

#7 sgt4boston

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Neutral

Posted 19 July 2013 - 11:18 AM

Try the different blocking suggestions and also prediluting your sample 1:10 in PBS.




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