Table of Contents
ISRN Urology
Volume 2012 (2012), Article ID 618247, 5 pages
http://dx.doi.org/10.5402/2012/618247
Research Article

Nitrotyrosine Density of Rabbit Urinary Bladder Muscle and Mucosa Measured via Western Blotting and 96-Well Plate Analysis

1Department of Pharmaceutical Sciences, Albany College of Pharmacy and Health Sciences, 106 New Scotland Ave, Albany, NY 12208, USA
2Department of Research, Stratton VA Medical Center, 113 Holland Ave, Albany, NY 12208, USA
3Division of Urology, Albany Medical College, 25 Hackett Blvd, Albany, NY 12208, USA

Received 4 November 2011; Accepted 13 December 2011

Academic Editor: S. Yücel

Copyright © 2012 Brittany Fitzpatrick et al. This is an open access article distributed under the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

Abstract

Purpose. Nitrotyrosine was quantitated in rabbit bladder muscle and mucosa using two analytical systems: Western blotting analyses and a 96-well plate quantitative analysis kit. Materials and Methods. Rabbit bladder muscle and mucosa were obtained from control rabbits. For the Western analysis, the samples were loaded into a SDS page gel and then transferred to a PVDF membrane. The optical density was measured using a Kodak Scanner. Using the 96-well plate, the samples and standards were loaded, incubated with primary and secondary antibody, washed and vacuumed with 10x wash buffer three times between each incubation period. Stop buffer was added to the plate and the results were quantified via the plate reader. Results. For both muscle and mucosa tissue, the optical density readings were linear with tissue concentration; the concentration of nitrotyrosine in the mucosa was significantly higher than in the muscle. However, whereas the Western blot analysis is based on relative optical densities, the 96-well plate kit provides a truly quantitative analysis. Discussion. Mucosa tissue displayed a higher density of nitrotyrosine than did detrusor muscle tissue. This may well be due to the significantly higher metabolic activity of the mucosa compared to the muscle.