Figure S2.

Screening of optimal Cas9-RNP nucleofection protocols for KO in murine BMDCs. (A) Data from the initial optimization screen are shown in four heatmaps reporting the cell abundances and CD45 KO efficiency in CD24⁺ (left panels) and Sirpα⁺ (right panels) DCs. Red boxes indicate the five conditions that showed the highest KO efficiency while maintaining acceptable cell abundance. Data are from one experiment. (B) Confirmation of deletion efficiency of the top five conditions from the initial optimization using CD45 crRNAXT-Cas9-RNPs compared with NTC crRNAXT-Cas9-RNPs and no-nucleofection (NN) controls. Data are averaged from technical replicates. (C) Relative CD80 levels as measured by flow cytometry in BM-derived CD24⁺ DC, Sirpα⁺ DC, pDC, and macrophage cells not nucleofected (No Nuc) or nucleofected with NTC crRNAXT-Cas9-RNPs using either P3/CM-137 or P3/EN138 combinations. Data are presented as mean ± SEM (n = 5) and collected from two independent experiments.