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Human peripheral blood mononuclear cells (PBMCs) were stained with CD4 antibodies or with the corresponding REA Control (S) antibodies (left images) as well as with CD3 antibodies. Flow cytometry was performed using the MACSQuant®
Analyzer. The Tandem Signal Enhancer has been used to increase binding specificity of tandem-dye–conjugated antibodies. Cell debris and dead cells were excluded from the analysis based on scatter signals and propidium iodide fluorescence or 4',6-diamidino-2-phenylindole (DAPI) fluorescence, as in the case of tandem conjugates.
In order to compare the epitope specificity of REAfinity Clone REA623 with other known clones, a competition assay was performed.
Cells were incubated with an excess of purified unconjugated REAfinity Antibody followed by staining with fluorochrome-conjugated antibodies of other known clones against the same marker. Based on the fluorescence signal obtained, the clones were identified as recognizing completely overlapping (++), partially overlapping (+), or completely different epitopes (-) of the marker.
|Other clones||Overlap in epitope recognition with REA623|