Influence of Manual Double-Spin Preparation Protocols on the Cellular and Growth Factor Composition of Equine Platelet-Rich Plasma

Abstract

This prospective, randomised, ex vivo study investigated the effects of manual double-spin centrifugation protocols on the cellular and growth factor composition of equine platelet-rich plasma (PRP). Blood from 20 healthy horses was evaluated in two phases. In Phase A (n = 10), nine first-spin protocols were compared, and two were selected based on platelet enrichment and leukocyte reduction. In Phase B (n = 10), these were combined with nine second-spin conditions, generating 18 PRP preparations. Platelet (PLT), leukocyte (WBC) and erythrocyte (RBC) concentrations and PDGF-BB and TGF-β1 concentrations were measured. No significant differences in cellular composition were identified among first-spin protocols (p > 0.05). From Phase B, all final preparations achieved platelet enrichment and leukocyte depletion, with negligible RBC contamination. Second-spin force significantly affected PDGF-BB and TGF-β1 concentrations (p < 0.001). PDGF-BB increased from 2629 pg/mL (95% CI: 2111–3274) at 200× g to 3467 pg/mL (95% CI: 2784–4317) at 1000× g, while TGF-β1 increased from 13,364 pg/mL (95% CI: 11,286–15,824) to 16,998 pg/mL (95% CI: 14,355–20,127). TGF-β1 also increased with second-spin duration (p = 0.01). Platelet concentration was positively correlated with PDGF-BB (ρ = 0.604) and TGF-β1 (ρ = 0.691; both p < 0.001), whereas WBC concentration showed no significant association with either growth factor. Thus, both preparation methodology and donor-related variability influence equine PRP composition, indicating that protocol standardisation alone is unlikely to ensure biologically consistent PRP products.