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Hi Everybody,
I try to distinguish in rats endogenous and exogenous insulin
Can somebody help me?
David
FRANCE
[Radio-labeled? - db]
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Not sure about separation with HPLC but if you are testing human insulin in
rats you might want to look at an RIA or ELISA that does not react with rat
insulin, there is one available through Linco I think.
Jim
James D. Talton, Ph.D.
President & CEO
Nanotherapeutics, Inc.
12085 Research Drive, Suite N
Alachua, FL 32615
www.nanotherapeutics.com
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I am not familiar with an HPLC method to distinguish endogenous from
exogenous insulin.
But you might consider assaying sample for C-peptide (ELISA kit) since
it is indicator of endogenous insulin. Note: C-peptide is different and
distinct from C-reactive peptide/protein.
C-peptide is the connecting peptide between A- and B-chains of insulin
that is cleaved and released with endogenous insulin from beta cells of
pancreas. It is secreted with insulin in equimolar ratio, one mole of
C-peptide for each mole of insulin. It is useful for assessing diabetic
patients ability to produce insulin even if they are taking exogenous
insulin. But be aware that C-peptide has lower hepatic clearance
compared to insulin and higher urinary clearance than insulin, so
depending on site of blood collection, C-peptide levels may not reflect
molar equivalence of insulin at that site.
Exogenous insulin does not contain C-peptide.
Hope this helps,
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David
Two possible solutions come to mind:
1. C-peptide. Endogenous insulin is co-secreted with C-peptide (in
one-to-one molar ratio). Based on C-peptide kinetics, you will information
about prehepatic insulin secretion rate but this involves deconvolution.
2. Specific assay. Some insulin analogues such as Lispro have specific
insulin assays (have seen advertisements for such an assay but have never
used). Standard assay will give total insulin concentration, specific will
give Lispro (exogenous insulin) concentration. This assumes that you are
able to use one of these short acting insulin analogues.
Roman
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Unless you label the material with a radio, fluorescent or UV dye you
cannot
distinguish. The use of a biomarker may help, but a labelled probe is
the
best approach. You should also consider before and after dosing. Ed
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Copyright 1995-2010 David W. A. Bourne (david@boomer.org)