A Colorimetric Assay for Glycoproteins Based on the Periodic Acid/Schiff Stain

Abstract

of trypsin without EDTA. A monolayer of radioactively labelled BHK cells was incubated at 37°C for2min in 0.25 % trypsin; one half of the cells in suspension were harvested, washed and assayed for adhesion, and the other half of the cells were incubated for a further 15min in 0.25% trypsin before harvesting, washing and assaying for adhesion. Cells incubated in trypsin for only 2min did not show a Mn2+-stimulated adhesion that was much greater than the CaZ+/Mg2+-stimulated adhesion. In contrast, BHK cells incubated for 15min in trypsin showed an Mn'+-stimulated adhesion that was twice that for the Ca2+/Mg2+-stimulated adhesion. These cells completely excluded Trypan Blue. When such strongly trypsin-treated '4C-labelled BHK cells were preincubated in complete growth medium for up to 60min before assaying for adhesion, this difference between adhesion in Mn2+ buffer and Ca2+/MgZ+ buffer decreased. Thus the CaZ+/Mg2+-stimulated adhesion was 51 and 86% respectively of the MnZ+stimulated adhesion for BHK cells that had been freshly treated with trypsin and for the same cells that had been allowed to recover for 60min in growth medium. MnZ+ is considered to affect the cytoskeleton of the cell, possibly by its release of membrane-bound Ca2+ (Rabinowitch & DeStefano, 1973; Gwynn et al., 1976). Our experiments show, firstly, that Mn2+ causes abnormal spreading and does not replace the requirement for CaZ+/Mg2+ and secondly, that changes that occur with cell transformation or proteolysis for trypsin potentiate this atypical MnZ+ effect.

Periodic acid–Schiff’s reagent assay for carbohydrates in a microtiter plate format

“…The protocol was optimised using 25 L of GalNAc solution per well (1 mg/ml, approximately 113 nmoles) and was initially based on the test tube method of Mantle [18].…”

The effect of dietary sainfoin (Onobrychis viciifolia) on local cellular responses toTrichostrongylus colubriformisin sheep

“…Mast cells and eosinophils were enumerated following staining with toluidine blue (Enerback, 1966) and carbol-chromotrope (Lendrum, 1944), respectively. Goblet cells in the epithelium and paneth cells in the crypts were detected using Periodic acidSchiffs (PAS) (Mantle and Allen, 1978). Neutral and acidic mucins were identified through alcian blue-PAS staining (Newlands et al 1990).…”

Pic, an Autotransporter Protein Secreted by Different Pathogens in the Enterobacteriaceae Family, Is a Potent Mucus Secretagogue

“…To determine glycoprotein concentrations in samples from rat intestinal mucus, a colorimetric method was used (29). Schiff reagent was prepared by dissolving 1 g basic fucsin in 100 ml of water (at 100°C).…”