We overcome the issue of maintaining non-adherent hematological cells by using cytocentrifugation, and optimized buffers, incubation times, and washing steps. Because hematological cells are more resistant to permeabilization and to preserve protein integrity, we increased permeabilization time to 1 h 20 at 4°C, instead of 30 min at 37°C as recommended by the manufacturer. We adapted the primary antibody buffer to our cells. We increased all washing times and the stringency of buffers to reduce non-specific fluorescence. We increased the ligation incubation time from 30 min to 1 h. Finally, we optimized the amplification step (80 min at 37 °C instead of 100 min) to reduce the potential coalescent signals.