ELISA question

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hanhan2008
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ELISA question

Post by hanhan2008 » Fri Apr 25, 2008 5:02 pm

I'm doing MIP-2 ELISA for a cell line using a kit from Biosource (now part of Invitrogen). Reproducibility is bad in terms of the equations obtained from the standard curves and the values obtained between different experiments. The big intercepts (big blank readings) caused big variations in calculated MIP-2 values.
Could anybody give me suggestions on how to get better results, including tricks to deal with 96-# plate? Thanks!

hanhan2008
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Post by hanhan2008 » Fri Apr 25, 2008 6:53 pm

Detailed questions:
1. Could ELISA kits of different batches make significant differences?
2. Which company provides the best ELISA kits, or MIP-2 kits in particular? Or does the company matter?
3. If you always do ELISA the same way with little variations in incubation times and such, is it true that you always get quite similar equations from standards (slope and intercept)?
4. I found that for my MIP-2 ELISA, the standard curve is quite linear (0-250 pg/ml) with an R^2 of 0.98-0.99. Is a linear curve the usual case for ELISA in general?
5. In obtaining the equation from the standards, is the blank value used to get standard curve or we deduct the blank value directly from all OD readings? as we know, as long as R^2 is <1, usually the intercept doesn't equal the blank value.
Sorry for so many Qs but I cannot help myself out any time soon.
Thanks.

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