Paired indirect immunoenzyme staining with primary antibodies from the same species. Application of horseradish peroxidase and alkaline phosphatase as sequential labels

1984 ◽  
Vol 16 (5) ◽  
pp. 477-487 ◽  
Author(s):  
Kolbj�rn Valnes ◽  
Per Brandtzaeg
Keyword(s):  
Alkaline Phosphatase ◽  
Horseradish Peroxidase

scholarly journals New chromogens for alkaline phosphatase histochemistry: salmon and magenta phosphate are useful for single- and double-label immunohistochemistry.

1994 ◽  
Vol 42 (4) ◽  
pp. 551-554 ◽  
Author(s):  
C Avivi ◽  
O Rosen ◽  
R S Goldstein
Keyword(s):  
Alkaline Phosphatase ◽  
Monoclonal Antibodies ◽  
Molecular Biology ◽  
Horseradish Peroxidase ◽  
Reaction Products ◽  
Immunocytochemical Detection ◽  
Hematoxylin Staining ◽  
Double Label ◽  
Biology Research

Two new substrate chromogens for alkaline phosphatase (ALP) detection have been recently synthesized for use in molecular biology research, salmon and magenta phosphate. We show here that these two chromogens have advantageous characteristics for immunocytochemistry as well. Their relatively delicate pink- and magenta-colored products do not mask the colors produced by other staining procedures. In addition, the reaction products of these substrates are insoluble in water, ethanol, and xylene, permitting the use of regressive hematoxylin staining procedures and coverslipping in permanent resin-based media. Most importantly, when these ALP substrates are used in double-label immunocytochemistry in combination with horseradish peroxidase-diaminobenzidine (HRP-DAB) and counterstained with hematoxylin, all three colors can be easily distinguished. An application using these substrates for simultaneous immunocytochemical detection of two monoclonal antibodies of different classes, in combination with hematoxylin staining, is illustrated.

scholarly journals One-step double immunolabeling of mouse interdigitating reticular cells: simultaneous application of pre-formed complexes of monoclonal rat antibody M1-8 with horseradish peroxidase-linked anti-rat immunoglobulins and of monoclonal mouse anti-Ia antibody with alkaline phosphatase-coupled anti-mouse immunoglobulins.

1991 ◽  
Vol 39 (12) ◽  
pp. 1719-1723 ◽  
Author(s):  
T Krenács ◽  
H Uda ◽  
S Tanaka
Keyword(s):  
Alkaline Phosphatase ◽  
Horseradish Peroxidase ◽  
B Lymphocytes ◽  
Enzyme Reaction ◽  
Molecular Complexes ◽  
Simultaneous Application ◽  
One Step ◽  
Reticular Cells ◽  
Secondary Antibodies ◽  
Double Immunolabeling

A novel one-step double immunolabeling method was elaborated on the basis of the simultaneous application of preformed molecular complexes of two primary antibodies with their specific secondary antibodies labeled with different enzymes. Treatment with a rat monoclonal antibody (MAb), M1-8, pre-coupled with horseradish peroxidase-linked sheep anti-rat immunoglobulins, and enzyme reaction revealed by the 3-amino-9-ethylcarbazole/hydrogen peroxide reaction, resulted in red-brown intracytoplasmic staining of interdigitating reticular cells in the lymph nodes of Balb/c mice. Another molecular complex, made of mouse anti-Ia MAb with alkaline phosphatase-linked rabbit anti-mouse immunoglobulins, applied at the same time and then developed with naphthol AS-BI-phosphate/fast blue BB as substrate, yielded blue surface staining of this cell type in addition to labeling of B-lymphocytes. The method described provides the possibility of relatively rapid double antigen detection where the binding sites of the secondary antibodies are saturated by the specific primary immunoglobulins. This approach seems to avoid nonspecific binding of primary antibodies to Fc receptors, and the unwanted binding of secondary antibodies with cell surface immunoglobulins on B-lymphocytes or with crossreactive primary antibodies used in the other sequence, if the primary antibodies and the tissue are the same or crossreactive animal species.

Comparison of alkaline phosphatase and horseradish peroxidase conjugated antisera in the ELISA test for antibodies to reovirus 3, mouse hepatitis and Sendai viruses

1981 ◽  
Vol 15 (1) ◽  
pp. 69-74 ◽  
Author(s):  
P. Carthew ◽  
J. Gannon ◽  
I. Whisson
Keyword(s):  
Alkaline Phosphatase ◽  
Horseradish Peroxidase ◽  
Virus Type ◽  
Sendai Virus ◽  
Hepatitis Virus ◽  
Mouse Hepatitis Virus ◽  
Elisa Test ◽  
Serological Response

The method of enzyme-linked immunoadsorbent assay (ELISA) has been applied to the detection of antibodies to reovirus 3, Sendai virus and mouse hepatitis virus (type 1), and the serological response of mice after infection has been followed for 28 days to investigate the earliest appearance of ELISA titres. This has been compared to the appearance of haemagglutination-inhibiting and complement-fixing antibodies. Alkaline phosphatase conjugated antiserum produces the most sensitive and convenient ELISA for the murine viruses examined.

Naked-eye sensitive detection of alkaline phosphatase (ALP) and pyrophosphate (PPi) based on a horseradish peroxidase catalytic colorimetric system with Cu(ii)

The Analyst ◽  
2016 ◽  
Vol 141 (19) ◽  
pp. 5549-5554 ◽  
Author(s):  
Dongmin Shi ◽  
Yue Sun ◽  
Lin Lin ◽  
Chunjun Shi ◽  
Guangfeng Wang ◽  
...  
Keyword(s):  
Alkaline Phosphatase ◽  
Horseradish Peroxidase ◽  
Colorimetric Method ◽  
Sensitive Detection ◽  
Naked Eye ◽  
Colorimetric System ◽  
System P

In this paper, a novel colorimetric method for the detection of alkaline phosphatase (ALP) and pyrophosphate (PPi) was designed based on a Cu2+–horseradish peroxidase (HRP)–3,3′,5,5′-tetra-methylbenzidine (TMB)–H2O2 system.

scholarly journals Solid-phase enzyme immunoassay for immunoglobulin M antibodies to cytomegalovirus

1977 ◽  
Vol 5 (6) ◽  
pp. 629-634
Author(s):  
H Schmitz ◽  
H W Doerr ◽  
D Kampa ◽  
A Vogt
Keyword(s):  
Alkaline Phosphatase ◽  
Horseradish Peroxidase ◽  
Enzyme Immunoassay ◽  
Indirect Immunofluorescence ◽  
Solid Phase ◽  
Immunoglobulin M ◽  
Nuclear Antigen ◽  
Immunofluorescence Method ◽  
Infected Cells

A sensitive, solid-phase enzyme immunoassay for the detection of immunoglobulin M antibodies to cytomegalovirus is described. The results of the enzyme immunoassay correlated well with those obtained by an indirect immunofluorescence method. Horseradish peroxidase proved to be a more sensitive label than alkaline phosphatase. Nonspecific reactions, occurring with commercially available cytomegalovirus antigens, could be avoided by using a nuclear antigen prepared from sonically disrupted nuclei of cytomegalovirus-infected cells.

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