Repeated immunostaining of the same tissue section using alkaline phosphatase as a reporter
| Autor: | AA Smith |
|---|---|
| Rok vydání: | 2016 |
| Předmět: |
0301 basic medicine
Histology biology Staining and Labeling Chemistry Phosphatase General Medicine Alkaline Phosphatase Primary and secondary antibodies Molecular biology Stain Horseradish peroxidase Immunohistochemistry Staining 03 medical and health sciences Medical Laboratory Technology Sebaceous Glands 030104 developmental biology biology.protein Alkaline phosphatase Humans Immunostaining Skin |
| Zdroj: | Biotechnichistochemistry : official publication of the Biological Stain Commission. 91(6) |
| ISSN: | 1473-7760 |
| Popis: | One can determine the best dilution of a primary antibody for immunohistochemistry that uses horseradish peroxidase conjugated to a secondary antibody by testing increasing concentrations sequentially on the same tissue section. When the same tissue section is incubated repeatedly with increasing concentrations of primary antibodies to epithelial membrane antigen, smooth muscle α-actin, or vimentin using alkaline phosphatase conjugated to a secondary antibody as the reporter, the best staining was obtained with a less concentrated primary antibody than was optimal for a single staining test. The best concentration of primary antibody for single run staining using an alkaline phosphatase reporting system is usually four times the best concentration for staining with multiple runs. The optimal concentration can be determined by denaturing the residual alkaline phosphatase and extracting residual stain by incubating the section in 4:1 diglyme:phosphate buffered saline for 20 min at 80(o) C between tests of primary antibody concentrations. I tested the method for four chromogens from one supplier and one chromogen from a different supplier. |
| Databáze: | OpenAIRE |
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