ERp5-deficient platelets have enhanced activation of collagen signaling pathways, increased Ca²⁺ mobilization and aggregation in response to U44619, and increased adhesion to fibrinogen under shear. (A) Representative immunoblots of P-AKT and P-PLCβ3 in response to IIα stimulation. Band densitometry of P-AKT expressed as a ratio to glyceraldehyde-3-phosphate dehydrogenase (GAPDH); P-PLCβ3 expressed as a ratio to total protein in lysates of control ERp5^fl/fl (gray) and CKO Pf4Cre⁺/ERp5^fl/fl (red) platelets treated with buffer (light box) or IIα (dark box). (B) Representative immunoblots of P-SYK and P-SRC in response to CRP stimulation. Band densitometry of P-SYK and P-SRC expressed as a ratio to the total protein in lysates of control (gray) and ERp5 KO (red) platelets treated with buffer (light box) or CRP (dark box). (C) Representative immunoblots of P-p38 and p38 in response to stimulation with 0.2 μm U46619. Band densitometry of P-p38 expressed as a ratio to the total protein in lysates of control (gray) and ERp5 KO (red) platelets treated with buffer (light box) or U46619 (dark box). All ratios are presented as fold change over buffer-treated controls. (D) Calcium flux, representative image, and peak Ca²⁺ flux after stimulation of platelets from control ERp5^fl/fl (gray) or CKO Pf4Cre⁺/ERp5^fl/fl (red) mice with U46619. (E) Aggregation (%) after activation with ADP, thrombin, collagen, and U46619. (F) Representative images of platelets labeled with calcein after perfusion on fibrinogen-coated microfluidic channels at a shear rate of 500 s⁻¹ for 5 minutes. Kinetics of adhesion of washed platelets from control (gray) and CKO (red) mice. Area under the curve (AUC) of platelet surface fluorescence at the end of perfusion (300 seconds) for control (gray) and CKO (red) platelets. n = 3 to 4 per genotype for signaling proteins; n = 3 to 6 per genotype for Ca²⁺ flux, aggregation, and adhesion; mean ± SEM, Student t test (F), 1-way ANOVA with Dunn post hoc analysis (A-E). ∗P < .05.