Dear Andrew, I assume that you refer to antibodies-conjugated to PE-Cy5. At CLB we also developed monoclonal antibodies conjugated to PE-Cy5. During the development of these conjugates we have tested many commercially available PE-Cy5 conjugates. We found that the quality of the conjugates is very much depending of the company who produced them. Non-specific binding (especially with monocytes) is often seen if the conjugates are not optimally produced, purified and formulated. In some cases we have been able to reduce a-specific staining by diluting the antibody in PBS containing 1% BSA. In our own conjugates we use a special buffer formulation which prevent any remaining a-specific staining. Best regards, John Voorn Product manager CLB Reagents tel: +31-20-5123246 Plesmanlaan 125 fax: +311-20-5123570 1066CX Amsterdam E-mail: J_Voorn@CLB.nl <mailto:J_Voorn@CLB.nl> the Netherlands Site: WWW.CLB.nl <http://WWW.CLB.nl> -----Original Message----- From: adwells@mail.MED.UPENN.EDU [SMTP:adwells@mail.MED.UPENN.EDU] Sent: dinsdag 29 juni 1999 23:46 To: cytometry@flowcyt.cyto.purdue.edu Subject: Cy5 binding to cells Received: from ns-clb.clb.nl by ln210-2.clb.nl (Lotus SMTP MTA v1.05 (274.9 11-27-1996)) with SMTP id C12567A0.005D61BB; Wed, 30 Jun 1999 19:00:59 +0200 Received: from flowcyt.cyto.purdue.edu by ns-clb.clb.nl (AIX 4.1/UCB 5.64/4.03) id AA13782; Wed, 30 Jun 1999 20:07:22 +0200 Received: (from root@localhost) by flowcyt.cyto.purdue.edu (8.9.3/8.9.3) id KAA02678 for cyto-sendout; Wed, 30 Jun 1999 10:49:45 -0500 Received: (from daemon@localhost) by flowcyt.cyto.purdue.edu (8.9.3/8.9.3) id RAA31499 for kelley; Tue, 29 Jun 1999 17:32:27 -0500 Received: from mail.med.upenn.edu (MAIL.MED.UPENN.EDU [165.123.128.11]) by flowcyt.cyto.purdue.edu (8.9.3/8.9.3) with ESMTP id RAA31495 for <cytometry@flowcyt.cyto.purdue.edu>; Tue, 29 Jun 1999 17:32:26 -0500 Received: from [130.91.197.87] (RENAL32.MED.UPENN.EDU [130.91.197.87]) by mail.med.upenn.edu (8.9.3/8.9.3) with ESMTP id SAA10426 for <cytometry@flowcyt.cyto.purdue.edu>; Tue, 29 Jun 1999 18:45:46 -0400 (EDT) Message-Id: <v03102803b39efcb83bb6@[130.91.197.87]> Mime-Version: 1.0 Content-Type: text/plain; charset="us-ascii" Date: Tue, 29 Jun 1999 18:46:25 -0400 To: Cytometry Mailing List <cytometry@flowcyt.cyto.purdue.edu> From: "Andrew D. Wells, Ph.D." <adwells@mail.MED.UPENN.EDU> Subject: Cy5 binding to cells We have a problem with Cy5 binding (apparently) non-specifically to lymphocytes. Has anyone else seen this and are there any suggestions as to how to avoid this? Thanks, Andrew Andrew D. Wells, Ph.D. University of Pennsylvania Department of Medicine 904 Stellar-Chance Laboratories 422 Curie Boulevard Philadelphia, PA 19104 (215) 898-1951 (215) 573-2880 (FAX) adwells@mail.med.upenn.edu
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