FIG 2.

Syk-deficient neutrophils lack the ability to kill Candida species. (A) Western blotting of Syk expression comparing parental Cas9 neutrophils, primary bone marrow-derived PMNs, and Syk knockout ER-HoxB8 neutrophils generated using CRISPR-Cas9. (B) Wright-Giemsa staining of GMP myeloblasts, Syk KO cells, Cas9 parent cells, and primary bone marrow-derived neutrophils. (C) Cellular ATP levels of Cas9 parent cells, Syk KO PMNs, and primary bone marrow-derived PMNs, as measured by CellTiter-Glo assays. (D) Myeloperoxidase levels of Cas9 parent cells, Syk KO PMNs, and primary bone marrow-derived PMNs, as measured by TMB. (E) Surface expression levels of Ly6G and Mac-1 on Cas9 parent cells, Syk KO cells, and primary bone marrow-derived neutrophils, as measured by antibody staining and flow cytometry. (F) Cas9 and Syk KO neutrophil cell lines were coincubated with C. albicans, C. glabrata, and C. auris, at an MOI of 0.2, to assess killing levels. Yeast viability after coincubation and PMN lysis was determined by PrestoBlue fluorescence. *, P ≤ 0.05; **, P ≤ 0.01; ***, P ≤ 0.001; ****, P ≤ 0.0001; ns, not significant. Data represent results from a minimum of three independent experiments. ID, identifier.