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High-Sensitivity IHC Detection of Phosphorylated p27/Kip1 in Human Tissues Using Secondary Antibody Conjugated to Polymer-HRP

High-Sensitivity IHC Detection of Phosphorylated p27/Kip1 in Human Tissues Using Secondary Antibody Conjugated to Polymer-HRP

Michael Grahek, Ana Ptak, Alexander E Kalyuzhny

Methods Mol Biol. 2017;1554:211-218.

PMID: 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.

Chemicals Related in the Paper:

Catalog Number	Product Name	Structure	CAS Number	Price
IAR42417095	Anti-phospho-p27 Kip1 (pThr198) antibody produced in rabbit			Price
IAR4247025	Anti-Acylglycerol kinase antibody produced in goat			Price

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