Double immunofluorescence labeling for CD31 and CD105 as a marker for polyether polyurethane-induced angiogenesis in mice

Abstract

A crucial component of the integration between foreign implants and the host is angiogenesis. However, to date, none of the available techniques and/or endothelial markers employed to assess angiogenesis in the implant/host interface seems to be able to highlight vascular structures convincingly. In the present study we investigated and compared the expression of two endothelial cell markers: platelet endothelial cell adhesion molecule (PECAM-1) (CD31) and endoglin (CD105) using immunohistochemistry (IHC) and immunofluorescence (IF) to identify and quantify newly formed blood vessels in subcutaneous implants of polyether-polyurethane sponge of formalin-fixed paraffin-embedded tissue. At day 14 post implantation the discs of the synthetic matrix were removed and processed for histological and morphometric analysis. In IHC staining for CD31 antibody the number of vessels was 2.27±0.69 and 5.25±0.46 for CD105. In IF for CD31 the number of vessels was 15.36±1.295 and 10.54±0.8213 for CD105. The level of cross-reaction was lesser in IF images compared with IHC images. Colocalization of CD31/CD105 using confocal images showed positive correlation (Pearson's co-relation and Manders’ equation). The double labeling for blood vessels using the IF technique for CD31/CD105 may be an important tool for evaluation of angiogenesis in biomaterial/host integration.