Extraction of proteins from platelet alpha granules, for application onto chronic wounds to ameliorate the wound healing process

Abstract

Work done at these and other laboratories has shown that proteins like growth factors obtained from autologous platelets and applied to problematic wounds, was successful in hastening the wound healing process. The use of other platelet proteins, such as immunoglobulins, in this area is little known or investigated. This project presents a method of extracting immunoglobulin proteins from platelet alpha granules. The aim is to build up on the knowledge that has already been established, contributing in this way to the use of platelet proteins for wound healing. The method starts with the formulation of a platelet lysate from the platelets of healthy donors. Through this process, the platelets are broken down so that their alpha granules release the proteins contained therein. Once the lysates obtained, they are assayed for protein concentration values, after which each one undergoes a two-step process of High Performance Liquid Chromatography using a different column for each step. In the first step size-exclusion chromatography extracts albumin and immunoglobulin G. In the second step, a hydroxyapatite column, which is selective for IgG, extracts IgG. Once the proteins are obtained through HPLC, the result is confirmed through various techniques using electrophoresis, namely SDS-PAGE, Isoelectric Focusing and Flow Cytometry.