Background/Objectives: Monkeypox, twice declared a public health emergency of international concern by the WHO, currently lacks approved targeted therapeutics. This study focused on the development of monkeypox virus (MPXV) E8-specific human monoclonal antibodies (mAbs) derived from recipients of the recombinant vaccinia vaccine (rTV), with subsequent evaluation of their cross-neutralizing activity against orthopoxviruses, including the vaccinia virus (VACV) and MPXV. Methods: Three mAbs (C5, C9, and F8) were isolated from rTV vaccinees. Structural mapping characterized their binding domains on the MPXV E8 and VACV D8 proteins. Neutralization potency was assessed against the VACV TianTan strain and MPXV clade IIb. A combo was further evaluated in a VACV-infected mice model for clinical recovery and viral load reduction. Complement-dependent enhancement mechanisms were also investigated in vitro. Results: C9 targets the virion surface region of E8 and both the virion surface region and intravirion region of D8, showing cross-neutralization activity against the MPXV (IC50 = 3.0 μg/mL) and VACV (IC50 = 51.1 ng/mL) in vitro. All three antibodies demonstrated potent neutralization against the VACV in vitro: C5 (IC50 = 3.9 ng/mL), C9 (IC50 = 51.1 ng/mL), and F8 (IC50 = 101.1 ng/mL). Notably, complement enhanced neutralization against the VACV by >50-fold, although no enhancement was observed for the MPXV. In vivo administration accelerated clinical recovery by 24 h and achieved significant viral clearance (0.9-log reduction). Conclusions: E8-targeting mAbs exhibited broad-spectrum neutralization against orthopoxviruses, demonstrating therapeutic potential against both historical (VACV) and emerging (MPXV) pathogens. However, MPXV’s resistance to complement-dependent enhancement highlights the necessity for pathogen-adapted optimization. These findings establish E8 as a critical conserved target for pan-poxvirus VACV and MPXV countermeasure development.