Fig 6. Antigen-specific elimination of target cells by sTCR-Saporin as measured by flow cytometry.

(a) SupT1 cells expressing the indicated SCT were mixed at a 1:1 ratio with HLA-A2^neg SupT1 cells and co-cultured for 3 days in the presence of relevant or irrelevant sTCR conjugated to Saporin, as indicated. At end of culture, cells were stained with anti-HLA-A2 AF647 to determine the frequencies of HLA-A2 positive and negative target cells among FSC^hi,SSC^hi events using flow cytometry. (b) Assay was performed as described in (a) and the percentage of SupT1 cells expressing indicated SCT that was eliminated by addition of sTCR is shown, calculated as described in materials and methods. Error bars indicate SEM of n = 3. (c) HLA-A2^pos SupT1 cells mixed at a ratio of 1:1 with HLA-A2^neg SupT1 cells were incubated overnight with indicated concentrations of peptide, washed, and indicated sTCR-Sap was added. SupT1 cells expressing SCT were used as positive controls (disconnected symbols). Culture and analysis to determine the percentage of eliminated HLA-A2^pos targets was performed as described in (a,b). Results shown are representative of two experiments performed, and error bars represent SD of duplicates.