High-Sensitivity IHC Detection of Phosphorylated p27/Kip1 in Human Tissues Using Secondary Antibody Conjugated to Polymer-HRP.
Methods Mol Biol
; 1554: 211-218, 2017.
Article
em En
| MEDLINE
| ID: mdl-28185194
ABSTRACT
A complex composed of goat anti-rabbit secondary antibody conjugated to a polymer coated with horseradish peroxidase (HRP) molecules was used to develop rapid and highly sensitive immunostaining protocol for the detection of phosphorylated p27/Kip1 (T157) in human tissues. This polymer-HRP complex produced much better sensitivity detection compared to conventional biotin-streptavidin-HRP chemistry. Using polymer-HRP made it possible to reduce primary antibody concentration, eliminate some incubation steps such as avidin-biotin blocking and incubation with separate biotinylated secondary antibodies, and shorten the incubation time with primary antibody. Specificity of the detection was confirmed by eliminating labeling after treating tissues with lambda phosphatase to remove phosphate groups from p27/Kip1. Secondary antibodies conjugated to polymer-HRP is a reagent of choice in both research and diagnostic pathology allowing detecting low abundant and weakly expressed tissue targets.
Palavras-chave
Texto completo:
1
Base de dados:
MEDLINE
Assunto principal:
Fosfoproteínas
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Imuno-Histoquímica
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Imunoconjugados
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Anticorpos Fosfo-Específicos
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Inibidor de Quinase Dependente de Ciclina p27
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Peroxidase do Rábano Silvestre
Tipo de estudo:
Diagnostic_studies
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Guideline
Limite:
Humans
Idioma:
En
Ano de publicação:
2017
Tipo de documento:
Article