# Platelet-Rich Plasma (PRP) Before Clinical Application: Qualitative Flow Cytometric Analysis and Enzyme-Linked Immunosorbent Assay (ELISA) Exploring Platelet Activation and TGFβ Release During Storage

## TL;DR

This study examines how platelet-rich plasma (PRP) changes during preparation and storage, focusing on platelet activation and growth factor release to improve clinical applications.

## Contribution

The study provides new insights into platelet activation and TGFβ release dynamics during PRP preparation and storage.

## Key findings

- Platelet activation and degranulation increase progressively over time during PRP preparation and storage.
- TGFβ release in the supernatant peaks at 24 hours but decreases significantly after gelification.
- The PRP preparation protocol ensures a high-quality hemoderivative suitable for clinical use when applied at the right timing.

## Abstract

Background/Objectives: In clinical practice today, platelet concentrates are often used for topical surgical applications. They are biomaterials that can accelerate healing processes associated with oral and maxillofacial surgery as well as in several other clinical applications through the action of growth factors released by platelets at the surgical site. However, in most cases, the exact quantification of the released growth factors is challenging in both the short and long term. The aim of this study was to determine if early platelet activation and degranulation occur during the collection and utilization of platelet-rich plasma (PRP) in the surgical room, where, before its application, PRP undergoes a procedure of gelification via reactions with procoagulant agents. Methods: PRP was prepared from the blood samples of 39 patients following the modified Whitman protocol. The samples were then analyzed at four different time points (1, 6, and 24 h during preparation and clinical application in the surgery room) using flow cytometry and enzyme-linked immunosorbent assays (ELISAs) to investigate the platelet activation/degranulation and TGFβ release in the supernatant (SN) during storage and clinical application. The mean platelet count in the whole blood was 267.5 ± 48.58 × 103/mL (range: 189–334 × 103/mL), and the mean concentration was 2925.5 ± 833.37 × 103/mL (range: 748–3453 × 103/mL). Results: The activation and degranulation of platelet cells (measured via monoclonal antibodies: CD62p and CD63, respectively) demonstrated a progressive increase at 1 h, 6 h, 24 h, and after gelification. The TGFβ dosage in the supernatant (SN) at different times exhibited a similar trend, with a mean release of 18.36 ng/mL at 1 h, 21.96 ng/mL at 6 h, and 29.45 ng/mL at 24 h. After the gelification of the PRP, a significant reduction was observed, with a value of 15.52 ng/mL. Conclusions: The results reveal that the protocol used for the preparation, storage, and application of the PRP ensures a good-quality hemoderivative and that the platelet concentrate must be applied with the correct timing to support tissue healing processes.