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Dear friends :
I need to measure the concentration of endotoxin (LPS) in the plasma sample,
but it seems the test methods (LAL test) currently available ofen give
either false negative or false positive results due to some LAL reaction
inhibitors or enhancers contained in plasma. Could anyone kindly tell me how
to pretreat plasma sample to avoid the interference ? or is there any
reference or test kit I could look for ?
Thank you very much.
Jenny Zheng
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Unfortunantly, the measurement of endotoxin in serum remains
difficult. False positive occur for several reasons:
Endotoxin is present in the environment and contamination is
possible while handling the sample. Sterilization procedures
generally do not inactivate endotoxin. Therefore you will have
to use special treated or certified endotoxin-free blood
collection supplies. These are available commerically.
There are other substances in plasma that can cause activation of
LAL. These are controlled usually by sample dilution 1:10 to
1:20, although this procedure will reduce the sensitivity.
Centecor was using a capture antibody assay in which antibodies
to lipid A were coated on wells of a 96 well plate. The serum
was added to the wells, and incubated for a short period. Then
the wells were washed. LAL reagents were then added to the wells
to detect the bound endotoxin. I think that this method was
published in associated with an HA-1A publication.
False negative reactions also occur due to the presence of
inhibitors in serum. This problem can be dealth with in the same
manner as above. Harsh treatments such as addition of acid to the
same and heating >70C should be avoided since these can result in
loss of endotoxin.
Even with careful attention to procedures, it will be difficult to
validate the assay using standards employed for chemical assays in
terms of accuracy and precision.
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Try Contacting the Associates of Cape Cod. Their Phone number is (508)
540-3444.
Phillip Blatzheim
Baylor College of Medicine Phone: (713) 798-5190
Medicine -- Hypertension Fax: (713) 799-2469
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Copyright 1995-2010 David W. A. Bourne (david@boomer.org)