Overexpression of FVIII results in thrombosis. Either f8+/+;at3+/+ or f8–/–;at3–/– zebrafish, both in the fgb-gfp background, were incrossed and 1-cell stage embryos injected with pzf8, phF8-SQ, or phF8-ΔF and evaluated for spontaneous thrombi formation via fibrin deposition scoring at 5 dpf. pzf8 injected into (A) f8–/–;at3–/– and (D) f8+/+;at3+/+ zebrafish larvae exhibited statistically significant spontaneous thrombus formation. Empty vector demonstrates the baseline thrombus formation in f8–/–;at3–/– zebrafish larvae without functional FVIII activity. (B) phF8-SQ and (C) phF8-ΔF injected into f8–/–;at3–/– zebrafish larvae demonstrated statistically significant levels of spontaneous thrombi. Statistical significance was determined by ordinal logistic regression followed by Dunnett adjustments (∗∗∗P < .001; ∗∗P < .01).