Mickey Croyle croyleml at UC.EDU
Tue Jul 16 11:53:42 EST 1996

I=92m trying to get an EGF ELISA to work by following the method of G=
et al (Endocrinology, 135:  1854-1862, 1994).  This is an
antigen-inhibited ELISA method.  Mouse recombinant EGF is used to abs=
to the wells and as my standard.  I have worked with 2 polyclonal
antibodies.  The first was Harlan rabbit anti-EGF which was targetted
against the entire EGF protein.  The antibody I am currently working
with is Santa Cruz Biotechnology polyclonal Goat anti-mouse EGF in wh=
the antibody is specific for the first 19 amino acids of the
amino-terminus end of EGF.  5% BSA is used as the blocking agent.

=09I am interested in determining EGF levels in mouse blood and plasm=
a.  I
have been able to generate nice standard curves, but the problem aris=
when assaying the blood or plasma samples.  I test a series of 2:1
dilutions of the samples but the results do not show EGF levels
following the 2:1 pattern as expected.  Does this sound like an antib=
specificity problem or a need to =93clean up=94 the blood or plasma b=
running them through an ELISA?  I am still working out the appropriat=
dilution range for the Santa Cruz anitibody so the specificity questi=
is still open.

Thanks in advance.

Christopher Erwin
Pediatric Surgery Research
Children=92s Hospital Medical Center
3333 Burnet Ave.
Cincinnati, OH  45229-3026

Phone:  (513) 559-4822
Fax:    (513) 559-7657
e-mail: Croyleml at uc.edu

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