Based on a systematic evaluation of post-thaw characteristics, this study categorizes boar sperm into two distinct cryo-survival phenotypes: cryo-tolerant and cryo-sensitive. The freeze-tolerant group exhibited significantly superior post-thaw quality, including better motility, plasma membrane integrity, acrosome integrity, DNA integrity, and mitochondrial activity. Furthermore, these sperm demonstrated stronger antioxidant capacity and reduced oxidative damage (lower MDA levels). Through proteomic analysis, we identified seven key proteins associated with cold tolerance (e.g., HSPA12A, PRDX4). Functional validation revealed that supplementing the freezing extender with 1 µg/ml recombinant NT5C1B protein significantly improved multiple quality parameters in freeze-sensitive sperm, confirming its role as an effective factor in enhancing cryo-survival. In contrast, ADA protein supplementation showed no significant improvement. The well-established freezing protocol developed in this study provides additional options for boar semen cryopreservation.