I only got 2 responses but I thought a report back was due. 1. Reference #2 below cites that their cocktail of 18-22 Vbeta mAbs recognizes 70-75% of all peripheral blood T cells. 2. Your mileage may vary. 3 & 4. I was very impressed with the Coulter IOTest Beta Mark, at least in theory. It is an 8 tube assay which by a simple trick allows the determination of 24 TCR specificities. One Vbeta antibody is coupled FITC, another one PE, and a third specificity is a mixture of the FITC conjugate and the PE conjugate. Since allelic exclusion does not allow 2 TCRs to coexpress, you can use the double positives as a 3rd specificity. This still leaves FL3 and FL4 open for tabletop FACS users. Here is the URL with a description, but not much there: http://www.beckmancoulter.com/coulter/Cytometry/updates_sept00/newspecificit ies.asp I can send you a more detailed product description if you are interested. Just email me. 5. McCoy JP, Overton WR, Schroeder K, et al: Immunophenotypic Analysis of the T Cell Receptor VB Repertoire in CD4+ and CD8+ Lymphocytes from Normal Peripheral Blood. Cytometry 26:148-153, 1996. Van den Beemd R, Boor PPC, van Lochem EG, et al: Flow Cytometric Analysis of the VB Repertoire in Healthy Controls. Cytometry 40:336-345, 2000. > _______________________ > Calman Prussin > Laboratory of Allergic Diseases > NIAID/ National Institutes of Health > calman@nih.gov > > ---------- > From: Calman Prussin > Sent: Friday, March,30, 2001 10:34 > To: 'Cytometry Mailing List' > Subject: human TCR Vbeta typing > > I would like to TCR type a population of human antigen specific CD4 T > cells detected using intracellular cytokine staining. I am staining after > fixation, under which conditions I find 90-95% of mAbs "work". > > Questions: > > 1. Using commercially available anti-Vbeta mAbs, what % of the TCR > repertoire is "covered". > > 2. Is this type of analysis realistic or an experimental black hole? > > 3. Any suggestions for anti-Vbeta mAbs, particularly panels that have been > worked out? > > 4. Any suggestions for "tricks" to make this easier? One maneuver I am > considering is to pool a group of 4 anti-TCR mAbs for each initial > screening. That way I could cover the available repertoire in 4-5 tubes. > In those that are positive I would then go back and look at the 4 Vbetas > individually. > > 5. References? > > > Thanks, > > Calman >
This archive was generated by hypermail 2b29 : Sun Jan 05 2003 - 19:01:16 EST