ELISA assay in triplicate
Posted 11 February 2009 - 11:53 AM
Posted 12 February 2009 - 02:03 AM
I have been working with a commercially available ELISA kit that is known to have some problems. I do the samples in triplicate and it seems that when I look at the data there is one of the three results that does not agree with the other two. Can I disregard this data point?
Is possible that you have an edge effect between triplicates. This can be solved incubating your ELISA plates in an approppiate moist chamber.
You can read the Edge Well Effects from the link ELISA guide.
This phenomenon refers to the observation that occasionally ELISA results show a variance in their ODs between edge
wells and the wells in the central region of the plate. In the past this has been attributed to manufacturing variations in
plates leading to higher adsorption in the outer wells. More recently manufacturers have begun demonstrating and certifying
the consistency of the manufacturing process by testing and reporting the well-to-well variation of replicates
performed in all wells of a plate. For instance, Nunc certified plates are guaranteed to have a CV in replicate wells of <5%
and no wells with >10% variation.
Posted 12 February 2009 - 05:16 AM
Posted 12 February 2009 - 07:02 AM
Can I disregard this data point?
Your setup is to do samples in triplicate, then use the results unless there is a good specific reason to disregard a datapoint, just a difference in the results is not enough.
Pluralitas non est ponenda sine necessitate
-- "You must assume no plural without necessity".
Posted 12 February 2009 - 07:24 AM
Not enough repetitions in my mind.