Abstract
Abstract Immuno-Laser Capture Microdissection (LCM) and Expression Microdissection (xMD) are techniques that rely on immunohistochemistry (IHC) to specifically define target cells for microdissection. While these techniques have been successfully applied to LCM and xMD, the full effects of IHC on the biomolecules extracted from stained samples are not well characterized. The goal of our study was to evaluate the integrity of DNA, RNA and protein biomolecules obtained from tissue sections after standard IHC or immunofluorescence (IF) techniques. Utilizing snap frozen human prostate and ovarian carcinoma specimens, as well as mouse liver samples, we evaluated biomolecule recovery from immunostained samples, including histone and pan-cytokeratins AE1/AE3 targeted antibodies. Standard horseradish peroxidase and diaminobenzidine (DAB) based IHC was used as well as Alexa Fluor 488 for indirect-IF. In general, biomolecule recovery was diminished in IHC stained tissues compared to standard hematoxylin and eosin (H&E) staining. RNA was affected the most and showed significantly reduced quantity and quality after staining. DNA was recoverable, but quantities were reduced when compared to the standard H&E. However, PCR amplification was possible with all stained samples studied. Protein analysis also demonstrated a reduction of quantity of the proteins recovered from IHC stained samples, with a variable effect depending on the protein analysis methodology. The results demonstrated that IHC techniques using DAB labeling significantly affect the extracted biomolecules. Improved staining protocols are required to address these concerns and to further the capabilities of these potentially powerful microdissection techniques. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 3986.
Published Version
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