GENE TRANSFER ASSAYS AND IMAGING

University Of Pittsburgh At Pittsburgh, Pittsburgh PA

Investigators

Linked publications & trials

Abstract

The studies in this program project all utilize modified herpes simplex virus (HSV)-based vectors to[unreadable] develop and/or test gene transfer as a method to deliver analgesic proteins. Three important issues will[unreadable] need to be addressed in order to fully evaluate the success of these studies. First we must be able to[unreadable] demonstrate the successful delivery of the HSV vectors to the intended tissues. Second, we must be able[unreadable] to assess the expression of the HSV transgenes at both the mRNA and protein levels. And third, we must[unreadable] be able to examine the biological effects of transgene expression. The aim of this core is to provide[unreadable] standardized biochemical, molecular biological, and histological techniques to the three research projects[unreadable] so that they may adequately address these concerns. Towards this aim this core will perform: 1) Realtime[unreadable] quantitative PCR and reverse transcription PCR to measure vector load and transgene expression; 2)[unreadable] In situ hybridization (ISH) and immunocytochemistry (ICC) to localize HSV vectors, assess transgene[unreadable] expression, and examine any changes in endogenous gene and protein expression; 3) Enzyme-linked[unreadable] immunosorbent assay (ELISA), radioimmunoassay (RIA), high performance liquid chromatography[unreadable] (HPLC), and protein biochemistry (including Western blot) to quantify changes in transgene or[unreadable] endogenous tissue proteins; and 4) Ca2+ influx assays to examine a cDNA library for genes that interact[unreadable] with the nociceptive-specific vanilloid receptor (VR1). By performing these various assays under the[unreadable] direction of a single core, we can increase quality control, decrease variation, and reduce costs across the[unreadable] entire program project. The core will be utilized by all projects.