Dot Blot Analysis of Rat Gastric Mucin Using Histochemical Staining Methods
✍ Scribed by Y. Goso; K. Hotta
- Publisher
- Elsevier Science
- Year
- 1994
- Tongue
- English
- Weight
- 470 KB
- Volume
- 223
- Category
- Article
- ISSN
- 0003-2697
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✦ Synopsis
Rat gastric mucins blotted on various membranes were detected using the periodate-Schiff staining method. The use of a polyvinylidene difluoride (PVDF) membrane gave the best results compared to nitrocellulose, nylon, positively charged nylon, and positively charged PVDF membranes. Alcian blue, high-iron diamine, galactose oxidase-cold thionin Schiff, and paradoxical concanavalin A stainings were also performed to detect mucins on PVDF membranes. Mucins were quantitatively detected by this analysis using each staining method with a range of detection from 0.02 to (1 \mu) g. Mucins extracted from rat gastric mucosa were detected by these dot blot assays at the position of void volume during Sepharose CL-4B chromatography. Mucins, after being purified with cesium trifluoroacetate centrifugation, were also mainly excluded from a Sepharose CL-2B column and detected by the dot blot assay using each staining method. In contrast, mucins after reduced alkylation were separated into two fractions during Sepharose CL-2B chromatography; one was excluded but the other was included in the column. Both fractions were detected using the periodate-Schiff method, but only the excluded fraction was stained using the paradoxical concanavalin A staining method. Thus, the dot blot assay using histochemical staining methods is useful for detecting individual mucins during gel chromatography. O 1994 Academic Press, Inc.