Immunohistochemistry

University Of Pennsylvania, Philadelphia PA

Investigators

Linked publications & trials

Abstract

Core B: The Immunohistochemstry (IHC) Core will provide characterization of the composition and architecture of vascular lesions (atherosclerosis, aortic aneurysms and femoral artery wire-induced injury) (Projects 1 &4), and ofthe temporal and spatial regulation of vascular gene expression by hemodynamic forces and hypercholesterolemia (Project 5) using standardized protocols. Services include (i) training personnel in individual projects to harvest and embed tissue appropriately for IHC analysis, (ii) cutting and mounting of cryostat sections in a manner that allows quantitative analysis of morphologic regions over defined areas of lesion, (iii) histochemical and immunohistochemical staining and counterstaining of serial sections, (iv) image analysis of processed tissue and training in quantitative image analysis of personnel in individual projects, (v) generation of and training in the generation of publication quality composites of images. In addition to already standardized protocols for the detection of for example, various infiltrating inflammatory cell types, foam cells, endothelial cells, vascular smooth muscle cells (at various stages of differentiation), cycling cells, apoptotic cells and extracellular matrix components, the core will develop optimized protocols for detection of additional antigens (such as PAF for platelet involvement), markers of oxidative responses and markers of the unfolded protein response as needed to support each project. The Core Director will participate up front in designing animal studies that will require IHC analysis to ensure that tissues from optimal numbers of animals per group and at optimal time points will be available for analysis, and in determining the appropriate panel of targets to be analysed to address specific hypotheses regarding mechanisms underlying lesion the development, composition and architecture of lesions. The Core Director will assist individuals from the various projects in interpreting IHC results. Core activities will utilize existing equipment including a Microm HM505 E cryostat, Leica Leitz DMRD upright microscope with bright field illumination, Nikon E 600 upright microscope with bright field illumination. Image Pro Plus image analysis software (Media Cybernetics, Silver Spring, MD), HP color laserjet 2550 LN and Canon Image Press Cl printers. Procedures for coordinated scheduling and for assuring equitable access to core services are in place. The histochemical and immunohistochemical analysis provided by the Core will provide additional approaches for quantification of the extent of lesion and will provide important insights into the mechanisms that modulate the extent of lesions as well as the properties of lesions that can impact outcome, i.e. more fibrotic versus inflammatory atherosclerotic lesions that may translate into vulnerability to plaque rupture.