In article <1994Sep23.234126.21738 at wmichgw> 93huebner at wmich.edu writes:
>If anyone has any tips on Cayman Chemical's acetylcholinesterase EIA's,
>please drop me a line via e-mail. Thanks in advance...Jim
My lab assays about 10-20,000 for eicosanoids using Cayman AChE-linked EIAs.
In general we have found them to be reliable and work as advertised. First
tip -- unless you have a damned good reason not to, follow their instructions.
Second tip -- always use QC standards from a third party. The internal
standards provided with the kits are in some cases considerably different from
what they are supossed to be -- up to 50%. We have confirmed this discrepancy
by HPLC analysis as well as comparison to standards provided by three
different companies, especially for PGE2.
Tip 3. If you have any problems, call them. Kirk Maxey, the president of
Cayman is a solid scientist who genuinely tries to help his customers. Most
of the people who work for him have proven pretty reasonable.
Otherwise, remember the EIAs for eicosanoids are agic -- to expect an
Ab to distinguish between molecules which may differ in the orientation of
only one H or double bond in a meager 20-carbon compound is inherently
unreasonable. That they do is miraculous -- however, cross reactivities need
to be carefully controlled. For example, while the PGE Ab has only about
0.03% cross-reactivity with arachidonic acid, if you use 30uM AA as a
substrate, then assay product w/o purification, that will result in AA being
detected as 9 pMol/ml (nearly 3ng/ml) of PGE!
Also -- the PGE assay is only about 1/10th as sensitive as it used to be.
Advanced Magnetics (they're called something new now) claims to have an EIA
that goes below 1 pg/well but I have no firsthand knowledge of it.
Oh, BTW, the cAMP assay is internally consistent, it just doesnt give the same
numbers for either stds or samples that we've been getting with an RIA -- off
by 200-300%.
Any other questions? feel free to e-mail.
Ronald Walenga, Ph.D.
Assoc Prof
Depts Pediatrics and Physiology
CWRU Med School
rxw13 at po.cwru.edu
