ELISA hormon assay

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Dr.Stein
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ELISA hormon assay

Post by Dr.Stein » Thu Mar 29, 2007 12:25 pm

Anyone could help me how to configure or convert the OD value to the hormone level (ng/mL or ng/dL)? :oops:

I used to work with RIA and the manual (insert) mentions how to calculate from CPM to the hormone level but I cannot find it in ELISA manual insert :?

Thank you in advance.
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Vagabond
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Post by Vagabond » Thu Mar 29, 2007 6:26 pm

Quickly you plot the OD reading for you standard (say on the Y axis) similar to RIA and then across the X axis you plot you concentrations.

Draw the line (curve) get a ruler or calculate the slope of the line and use it to calculate what the OD of you sample is. Follow that down to the X axis and there is you concentration.

Use excel or some other program will be much easier. (softmax pro works well also).
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Post by Dr.Stein » Fri Mar 30, 2007 1:55 am

From your explanation, it seems to me that the graphic would be a linear one with the formula: y = ax+b since you mentioned "ruler"? Yes I can get the "r^2" automatically with Excel or another LR analysis tool.

Now I need to know about the scale. RIA uses log-logit scale to put dots and then draw the linear graphic. What scale must I apply for ELISA?

So, could I say that the principle to determine the hormone level using ELISA is similar to RIA, which using extrapolation method after drawing the standard curve?
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Post by blcr11 » Sat Mar 31, 2007 5:00 pm

How well it fits a straight line depends on how linear the assy is. ELISA standard curves should look similar to RIA curves. Logit-log transformations are used to linearize the curve over the range of at least 10-90% saturation.

Here is a brief overview of ELISA from Millipore Corp.
http://www.chemicon.com/resource/ANT101/a2C.asp

and a formal paper (in the public domain from PubMed) comparing different models for analysis of standard curves
http://www.pubmedcentral.nih.gov/picren ... obtype=pdf

and even an example using Prism, though I'm not sure this is very helpful
http://www.graphpad.com/prism/learn/Non ... curves.pdf

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Post by Dr.Stein » Sun Apr 01, 2007 2:47 am

Thank you, friends. I kept trying to get references and finally I found out the "formula" to draw the graphic! Yay! And yes, it is not linear. The hormone level will be determined from extrapolation of Y axis (OD sample/OD blank * 100) against X axis (standard's hormone level). Woo hoo I made it ! I made it! :D
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