[Histonet] RE: MMP-1

[C.M. van der Loos]

   Dear Samantha,

   You  have  done already a lot to make this MMP-1 antibody work.  There
   are only a few things left that you may try from here on:
     * A new  titration  series  (1:20  - 1:60 - 1:180) with an overnight
       primary   antibody  incubation  at  4C.  Tissue  pretreatment:  no
       treatment,  HIER  with  EDTA  pH9.0,  HIER with citrate pH 6.0 and
       pronase
     * Test   some   different   anti-MMP1   antibodies   from  different
       sources! To  my  opinion Santa  Cruz  antibodies  are not the best
       guarantee  for optimal staining results. With respect to other MMP
       antibodies  (3 and 9) we have good staining results with LabVision
       reagents. Perhaps one of their anti-MMP-1 antibodies will work for
       you.
     * In   case   of   negative   staining,  you  need  perhaps  a  more
       sensitive/efficient   detection   system.  Tyramide  amplification
       techniques  can  be  helpful  in  this  respect  (Perkin&Elmer TSA
       NEL-700 kit or DakoCytomation CSA II kit).

   Good luck,

   Chris van der Loos, PhD
   Dept. of Pathology
   Academical Medical Center M2-230
   Meibergdreef 9
   NL-1105 AZ Amsterdam
   The Netherlands

   phone:  +31 20 5665631
   fax:    +31 20 6960389


   Date: Mon, 27 Jun 2005 15:34:30 -0600
   From: "Samantha McMurtrey" <SMcMurtrey <@t> UniPathLLC.com>
   Subject: [Histonet] MMP-1
   To: <histonet <@t> lists.utsouthwestern.edu>
   Message-ID: <auto-000289236292 <@t> fe.mail.megapathdsl.net>
   Content-Type: text/plain; charset="us-ascii"
   Hello all,
   I  am  having  a  lot of trouble getting any staining for MMP-1.  I am
   using
   Santa Cruz, catalog # sc-21731, lot # A1405. It's a mouse monoclonal
   antibody.  I have tried dilutions from 1:200 down to 1:25. I incubated
   the
   primary  antibody  for 30 minutes and the secondary for 30 minutes.  I
   have
   also  tried  using proteinase K, Target, EDTA, and a pH 9 retrieval. I
   am
   working  it up on formalin fixed paraffin embedded tissue. I am mostly
   using
   breast   tissue   that   has  atypical  ductal  hyperplasia  (per  the
   pathologist's
   request).   I  haven't  had any luck at all. I am getting no staining.
   Please
   help!
   Thank you,
   Samantha McMurt! rey, BS,