Soluble TSP does not bind SS-RBCs. Washed SS-RBCs were incubated with purified TSP (1.4 mg/mL) in the presence of 1 mmol/L Ca²⁺ for 30 minutes, 37°C. Treated RBCs were incubated with the nonblocking anti-TSP MoAb A4.1 (SS1 + TSP, SS2 + TSP), isotype-specific control MoAb MBC 45.7 (Control), or a positive control MoAb directed against glycophorin C (glycophorin C), followed by FITC-conjugated anti-mouse MoAb, and bound TSP detected by flow cytometry. The above data are representative of six separate experiments. (B) Washed SS-RBCs were incubated with purified soluble TSP (100 μg/mL) for 30 minutes at 37°C before performing the flow adhesion assay over immobilized TSP as described above (N = 3).