Comparison of Automated and Manual immunohistochemical, Staining of Bromodeoxyuridine (BrdU)-Labeled Pulmonary Epithelial Cells in Mice

Abstract

An automated method for 5-bromo-2′-deoxyuridine (BrdU) staining of mouse lung tissue with the Fisher Code-On series is described. BrdU labeling indices (LI) were statistically analyzed to determine the efficacy of automated vs manual staining to detect BrdU incorporation. In the automated method, which utilized capillary gap technology, tissue-tissue and tissue-non-tissue slide pairing were evaluated. The effects of varying trypsin concentrations and the maximum concentration for producing optimal automated BrdU staining were also determined. Comparisons were made between 10% neutral buffered formalin and modified B5 (mercuric chloride) fixed tissues. The automated and manual BrdU staining methods were comparable in overall quality. Analysis of variance procedures revealed no significant differences between the manual and automated procedures for LI in the treated and control animals. In the automated method, no significant difference in the intensity or pattern of staining was found in the variations of slide pairing. A 0.3 % trypsin concentration enhanced the intensity of nuclear staining compared to 0.006 % trypsin; however, the optimal trypsin concentration in which cellular integrity was maintained was determined to be 0.0375 % . Tissues preserved in modified B5 fixative had distorted cellularmorphology and decreased intensity of staining compared to tissues fixed in 10% neutral buffered formalin, after trypsin digestion. (The J Histotechnol 17:23, 1994)

Keywords:

• automated staining
• bromodeoxyuridine (BrdU)
• imrnunohistochemistry
• lung
• mice