Optimizing Platelet-Rich Plasma Gel Formation by Varying Time and Gravitational Forces During Centrifugation
Despite the increasing clinical use of topical platelet-rich plasma (PRP) to enhance tissue healing and regeneration, there is no properly standardized method of autologous PRP gel preparation. This study examined the effect of the centrifugation time and gravitational force (g) on the platelet recovery ratio of PRP and determined the most effective centrifugation conditions for preparing PRP. Two-step centrifugations for preparing PRP were used in 39 subjects who had consented prior to the study's start. The separating centrifugation (Step 1, used to separate whole blood into its two main components: red blood cells and plasma) was tested from 500g to 1900g at 200g increments for 5 minutes (min), and from 100g to 1300g at 200g increments for 10 minutes. After separating centrifugation, upper plasma layer was transferred to another plain tube for the condensation centrifugation and remaining lower cell layer was discarded. The condensation centrifugation (Step 2, used to condense the platelets in the separated plasma) was tested at 1000g for 15 min, 1500g for 15 min, 2000g for 5 min and 3000g for 5 min, additionally at 1000g for 10 min and 1500g for 10 min. Platelet gelation was induced by adding 10% calcium gluconate to final PRP with volume ratio of 1:10. The optimal separating centrifugation conditions were followed by 900g for 5 minutes and the condensation conditions were followed by 1500g for 15 minutes, of which recovery ratios were 92.0 ± 3.1% and 84.3 ± 10.0%, respectively.
Automated cell count machine used in present study.
Figure 2. The separating centrifugation (Step 1). The erythrocytes are sedimented but platelets remain in suspension. (a) Initial anticoagulated blood. (b) After separating centrifugation. (c) A separated red blood cell layer. (d) The upper plasma layer was transferred to another plain tube. Figure 3. The condensation centrifugation (Step 2). (a) Plasma layer before condensation centrifugation. (b) Plasma layer after condensation centrifugation. (c) Platelet pellet after condensation centrifugation. (d) Supernatant layer (platelet-poor plasma) was discarded and lower 2 mL platelet-rich plasma was preserved. Figure 4. Gelling test. (a) Final 2 mL PRP, in which the platelet pellet was resuspended. (b) Calcium gluconate (0.2 mL, 10%) was added to final 2 mL PRP sample. (c) Platelet gel at 1 hour. (d) Platelet gel at 24 hours.
Figure 5. Histogram of platelet concentration of anticoagulated blood samples with 1.25 mL CPDA–1. (n = 39). CPDA–1 = citrate phosphate dextrose adenine–1. Figure 6. Platelet recovery ratios following separating centrifugation of 5 minutes. (Number of samples in each group = 5.) Significant difference (P < .05) between centrifugation conditions (Kruskall-Wallis test). c > a; c > h (P < .001, P = .001); d > a; d > h (P = .002, P = .006). Figure 7. Platelet recovery ratios following separating centrifugation of 10 minutes. (Number of samples in each group = 4.) Significant difference (P < .05) between centrifugation conditions (Kruskall-Wallis test). c > a; c > f; c > g (P = .001, P = .022, P < .001); b > a; b > g (P = .005, P = .003). Figure 8. Volume of platelet gel (mL) with time after adding 0.2 mL 10% calcium gluconate to 2 mL platelet-rich plasma (n = 6).
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