Expression of NBEAL2, SEC22B, and α-granule proteins in imMKCL WT, KO, and rescue cells. (A) Immunoblot analysis confirmed loss of NBEAL2 or SEC22B in CRISPR/Cas9 gene-edited NBEAL2-KO and SEC22B-KO cell lines and confirmed expression of HA-NBEAL2 and HA-SEC22B in the rescue lines NBEAL2-R and SEC22B-R. (B) Immunoblot analysis showed reduced expression of von Willebrand factor (VWF), thrombospondin-1 (TSP-1), and P-selectin in NBEAL2-KO and SEC22B-KO cells and restoration of expression of these proteins in NBEAL2-R and SEC22B-R cells. (C) Densitometric analysis of immunoblots indicated SEC22B-KO cells had significantly decreased NBEAL2 expression relative to WT (39.2% ± 11.8%), whereas NBEAL2-KO cells had normal levels of SEC22B (119.8% ± 14.0%). Relative protein levels represent the ratio relative to glyceraldehyde-3-phosphate dehydrogenase (GAPDH) normalized to the WT ratio. Data are representative of 3 independent experiments (bars = 95% confidence intervals). *P < .05 via 2-way analysis of variance with Dunnett’s post hoc test relative to WT.