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Diaminobenzadine

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Diaminobenzadine is a chemical compound used as a chromogenic substrate in various laboratory applications. It serves as a reagent for the detection and visualization of certain enzymes, particularly horseradish peroxidase, in immunohistochemistry and enzyme-linked immunosorbent assays (ELISA).

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Lab products found in correlation

Anti rabbit secondary antibody, by Merck Group (1 mentions) Biotinylated goat anti mouse rabbit igg, by Agilent Technologies (1 mentions) Biotinyted anti rabbit igg secondary antibody, by Vector Laboratories (1 mentions) Ncl ki67p, by Leica (1 mentions) Avidin biotin complex, by Vector Laboratories (1 mentions)

5 protocols using diaminobenzadine

1

Immunohistochemical Staining Protocol

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The sections were de-paraffinized and boiled in 10 mM citrate buffer (pH 6.0) for antigen retrieval. Endogenous peroxidase was blocked by 3% H2O2. Slides were blocked in serum, incubated with the indicated antibodies at 4°C overnight, incubated with anti-rabbit secondary antibody, and visualized with diaminobenzadine (Sigma). A negative control experiment was also performed. IHC staining images were captured at 200× under a microscope (Olympus).
Gao P., Wang H., Yu J., Zhang J., Yang Z., Liu M., Niu Y., Wei X., Wang W., Li H., Wang Y, & Sun G. (2018). miR-3607-3p suppresses non-small cell lung cancer (NSCLC) by targeting TGFBR1 and CCNE2. PLoS Genetics, 14(12), e1007790.
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2

Immunohistochemical Staining Protocol

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The section was de-paraffinized and boiled in 10 mM citrate buffer (pH 6.0) for antigen retrieval. Endogenous peroxidase was blocked by 3% H2O2. Then slides were blocked in serum, incubated with the indicated antibodies at 4 °C overnight, then incubated with anti-rabbit secondary antibody, and visualized with diaminobenzadine (Sigma). Negative control was also performed. The images of IHC staining were photographed at 200 × with microscope (Olympus).
Zhou Q., Zhu Y., Wei X., Zhou J., Chang L., Sui H., Han Y., Piao D., Sha R, & Bai Y. (2016). MiR-590-5p inhibits colorectal cancer angiogenesis and metastasis by regulating nuclear factor 90/vascular endothelial growth factor A axis. Cell Death & Disease, 7(10), e2413-.
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3

Immunohistochemical Detection of T. cruzi

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Hearts were fixed in buffered formalin solution, embedded in paraffin, and cut into 5 μm sections. Sections were stained with haematoxylin-eosin or used for the detection of T. cruzi antigens by means of the immunoperoxidase method. For the latter use, sections were deparaffinised and incubated with polyclonal anti-T. cruzi antiserum obtained from rabbits immunized with T. cruzi whole homogenate. Biotinylated goat anti–mouse/rabbit IgG and Duet Strepto ABComplex (Dako, Denmark) were used as a secondary antibody and for amplification, respectively. Diaminobenzadine (SIGMA Chemical Co, USA) was used as chromogen. preimmune rabbit serum was used as a negative control [17 (link)].
Navarro I.C., Ferreira F.M., Nakaya H.I., Baron M.A., Vilar-Pereira G., Pereira I.R., Silva A.M., Real J.M., De Brito T., Chevillard C., Lannes-Vieira J., Kalil J., Cunha-Neto E, & Ferreira L.R. (2015). MicroRNA Transcriptome Profiling in Heart of Trypanosoma cruzi-Infected Mice: Parasitological and Cardiological Outcomes. PLoS Neglected Tropical Diseases, 9(6), e0003828.
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4

Immunohistochemical Analysis of Phosphor-ERK and Ki67

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Immunohistochemical staining was performed using avidin-biotin-peroxidase complex method on paraffin-embedded sections as previously described. The primary antibodies are phosphor-ERK antibody (1:100) and Ki67 antibody (Rabbit mAb, 1:100 diluted, Vector Laboratories Inc, Burlingame, CA). Biotinyted anti-rabbit IgG secondary antibody (1:200) and ABC complex (Avidin-Biotin-Complex) were purchased from Vector Laboratories Inc, Burlingame, CA. Diaminobenzadine (Sigma-Aldrich, St.Louis, MO) was used as the chromogen. Slides were washed with 1XTBST buffer between incubations and counterstained with Mayer’s hemotoxylin for 1 minute. The number of phosphor-ERK positive-stained cells per high power (40× objective lens) was counted for each specimen. Cell proliferation was analyzed by Ki67-labeled cells and proliferation index was the percentage of Ki67 positive cells in the total number of cells counted.
Zhang W., Li H., Yang Y., Liao J, & Yang G.Y. (2014). Knockdown or inhibition of Aldo-keto reductase 1B10 inhibits pancreatic carcinoma growth via modulating Kras - E-cadherin pathway. Cancer letters, 355(2), 273-280.
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5

Hippocampal Neurogenesis Immunohistochemistry

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Immunohistochemical analysis was conducted using 40 μm thick sections for every 6th sagittal section of the right hemisphere. In order to ensure sampling of the entire hippocampus, the sections collected included those immediately medial and lateral of the hippocampus. Free floating sections were treated with 3% hydrogen peroxide in methanol to remove endogenous peroxidase activity. They were blocked with 10% normal serum corresponding to the species each secondary antibody was raised in (horse or goat) with 0.1% Triton X-100 diluted in PBS. Primary antibodies to doublecortin (DCX; Santa Cruz, SC-8066; 1:150) or Ki67 (Novocastra, NCL#Ki67p; 1:500) were also diluted with 3% serum with 0.1% Trition X-100 overnight, oscillating at 60 rpm at 4°. Secondary biotinylated antibodies were diluted 1:500 and 1:1000 in PBS containing 3% normal serum with 0.1% Triton X-100 for DCX and Ki67, respectively, and incubated for 2 h at room temperature. Avidin-biotin complex was used to amplify signal (Vector Labs), and diaminobenzadine (Sigma) was used for color development.
Winter A.N., Subbarayan M.S., Grimmig B., Weesner J.A., Moss L., Peters M., Weeber E., Nash K, & Bickford P.C. (2020). Two forms of CX3CL1 display differential activity and rescue cognitive deficits in CX3CL1 knockout mice. Journal of Neuroinflammation, 17, 157.
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