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3 3 diaminobenzidine dab chromagen

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3,3′-diaminobenzidine (DAB) chromagen is a chemical compound used as a chromogenic substrate in various immunohistochemical and enzymatic detection techniques. It serves as a sensitive and reliable method for visualizing specific target proteins or enzymes in biological samples.

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Lab products found in correlation

Hematoxylin, by Thermo Fisher Scientific (2 mentions) Permount, by Thermo Fisher Scientific (2 mentions) Hematoxylin counterstain, by Vector Laboratories (2 mentions) Anti mouse or anti rabbit secondary antibody, by Biocare Medical (2 mentions) Irrelevant isotype matched antibodies, by Biocare Medical (2 mentions) Adamts5, by Abcam (1 mentions) Proteinase k solution, by Thermo Fisher Scientific (1 mentions) Avidin biotin peroxidase complex, by Vector Laboratories (1 mentions) Mmp13, by Abcam (1 mentions) Mmp 1, by Abcam (1 mentions) Cited2, by Abcam (1 mentions)

Spelling variants of this product

DAB (673 citations) Diaminobenzidine (DAB) (96 citations) DAB (3,3′-Diaminobenzidine) (18 citations) DAB chromagen (6 citations)

5 protocols using 3 3 diaminobenzidine dab chromagen

1

Immunohistochemical Staining of Paraffin-Embedded Samples

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Paraffin-embedded blocks were cut in 5 μm sections. Slides were processed as follows: they were dewaxed in xylene followed by rehydration in a standard alcohol series. Antigen retrieval was done by pressure-cooking for 20 min in citrate buffer (pH 6.0), followed by blocking of endogenous peroxidase in 0.3% H2O2. 10% serum derived from the secondary antibody source was used to block for 30 min. Sections were incubated overnight at 4 °C with indicated primary antibodies. Antibodies were detected using a secondary HRP-labeled mouse or rabbit antibody detection system (Dako EnVision + System-HRP k4401, k4403) followed by addition 3,3′-diaminobenzidine (DAB) chromagen (Vector Labs) for visualization. Sections were counterstained with hematoxylin (Fisher Scientific Inc.) and slides were dehydrated in 70, 80, and 100% ethanol and xylene. Slides were covered by coverslips and mounted in Permount (Fisher Scientific Inc.). All images were captured on an Olympus IX73 fluorescence microscope system and analyzed using CellSens Dimension software.
Bunda S., Heir P., Metcalf J., Li A.S., Agnihotri S., Pusch S., Yasin M., Li M., Burrell K., Mansouri S., Singh O., Wilson M., Alamsahebpour A., Nejad R., Choi B., Kim D., von Deimling A., Zadeh G, & Aldape K. (2019). CIC protein instability contributes to tumorigenesis in glioblastoma. Nature Communications, 10, 661.
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2

Quantitative Immunohistochemical Analysis of Cartilage Degeneration

Cited 1 time
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Sections were incubated overnight at 4 °C with antibodies against cleaved aggrecan (NITEGE, Ibex) and cleaved type II collagen (Col2-3/4 M, Ibex), matrix metalloproteinase (MMP)-13 (Abcam), and a disintegrin and metalloproteinase with thrombospondin motifs (ADAMTS)5 (Abcam) followed by incubation with anti-mouse or anti-rabbit secondary antibody (Biocare Medical) and visualization with 3,3-diaminobenzidine (DAB) chromagen (Vector Laboratories). Negative controls were stained with irrelevant isotype-matched antibodies (Biocare Medical). Immunostaining intensity for type II collagen or aggrecan cleavage epitopes was quantified by determining the “reciprocal intensity” of the stained articular cartilage matrix; briefly, the light intensity value of six random locations within all three zones from the posterior to anterior direction of the femoral and tibial condyles of three sections per mouse was measured using the color picker in Adobe Photoshop [36 , 37 (link)]. Percentages of positive MMP-13 and ADAMTS5 chondrocytes were determined by counting the number of immunostained cells and dividing by the total number of chondrocytes visualized by a hematoxylin counterstain (Vector Laboratories).
Zhang Z., Leong D.J., Xu L., He Z., Wang A., Navati M., Kim S.J., Hirsh D.M., Hardin J.A., Cobelli N.J., Friedman J.M, & Sun H.B. (2016). Curcumin slows osteoarthritis progression and relieves osteoarthritis-associated pain symptoms in a post-traumatic osteoarthritis mouse model. Arthritis Research & Therapy, 18, 128.
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3

Immunohistochemical Localization of PTHR1

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PTHR1 was localized immunohistochemically using 3-μm sections from the same blocks used for histologic analysis. Sections were deparaffinized and rehydrated. Epitope retrieval was performed in a 0.1% proteinase K solution (Invitrogen, Carlsbad, CA) for 10 min at room temperature. The following primary antibodies were used: mouse monoclonal anti-PTH/PTHrP receptor (Upstate Cell Signaling Solutions, Charlottesville, VA, 1:100 dilution). Slides were treated with biotinylated secondary antibodies and developed with the avidin-biotin peroxidase complex and 3,3′-diaminobenzidine (DAB) chromagen (Vector Laboratories, Burlingame, CA) and then counterstained with hematoxylin.
Lee D.J., Southgate R.D., Farhat Y.M., Loiselle A.E., Hammert W.C., Awad H.A, & O'Keefe R.J. (2014). Parathyroid Hormone 1-34 Enhances Extracellular Matrix Deposition and Organization During Flexor Tendon Repair. Journal of orthopaedic research : official publication of the Orthopaedic Research Society, 33(1), 17-24.
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4

Immunohistochemistry Protocol for Tissue Sections

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Paraffin-embedded blocks were cut into 5-μm sections. Slides were processed as follows: de-waxed in xylene followed by rehydration in a standard alcohol series. Antigen retrieval was by pressure cooking for 20 min in citrate buffer (pH 6.0), followed by blocking of endogenous peroxidase in 3% H2O2. The antibodies were added and incubated overnight at 4 °C. Antibodies were detected using a secondary-HRP labeled mouse or rabbit antibody detection system (Dako EnVision+ System-HRP cat. no. k4401, cat. no. k4403) followed by addition of 3,3ʹ-diaminobenzidine (DAB) chromagen (Vector Labs) for visualization. Sections were counter-stained with hematoxylin (Fisher Scientific) and slides dehydrated in 70, 80 and 100% ethanol and xylene. Slides were cover-slipped and mounted in Permount (Fisher Scientific). IHC was performed as described by Koncar et. al.11 (link).
Golbourn B.J., Halbert M.E., Halligan K., Varadharajan S., Krug B., Mbah N.E., Kabir N., Stanton A.C., Locke A.L., Casillo S.M., Zhao Y., Sanders L.M., Cheney A., Mullett S.J., Chen A., Wassell M., Andren A., Perez J., Jane E.P., David Premkumar D.R., Koncar R.F., Mirhadi S., McCarl L.H., Chang Y.F., Wu Y.L., Gatesman T.A., Cruz A.F., Zapotocky M., Hu B., Kohanbash G., Wang X., Vartanian A., Moran M.F., Lieberman F., Amankulor N.M., Wendell S.G., Vaske O.M., Panigrahy A., Felker J., Bertrand K.C., Kleinman C.L., Rich J.N., Friedlander R.M., Broniscer A., Lyssiotis C., Jabado N., Pollack I.F., Mack S.C, & Agnihotri S. (2022). Loss of MAT2A compromises methionine metabolism and represents a vulnerability in H3K27M mutant glioma by modulating the epigenome. Nature cancer, 3(5), 629-648.
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5

Immunohistochemical Analysis of Hindlimb Chondrocytes

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Mouse hindlimbs were fixed in formalin, decalcified in formic acid, embedded in paraffin and sectioned for immunohistochemistry. Sections were incubated overnight at 4 C with antibodies against acetylated a-tubulin (Sigma), CITED2, MMP-1, and MMP-13 (all from Abcam), followed by incubation with AlexFluor 488 conjugated anti-mouse (for acetylated a-tubulin staining) or antimouse or anti-rabbit secondary antibody (Biocare Medical) and visualization with 3,3ʹ-Diaminobenzidine (DAB) chromagen (Vector Laboratories). Negative controls were stained with irrelevant isotype-matched antibodies (Biocare Medical). Cilia were counted by scoring acetylated a-tubulin-positive structures greater than 1 mm in length from at least five different views of each section 27 .
Percentages of positively stained chondrocytes were determined by counting the number of immunostained cells and dividing by the total number of chondrocytes visualized by a hematoxylin counterstain (Vector Laboratories).
He Z., Leong D.J., Zhuo Z., Majeska R.J., Cardoso L., Spray D.C., Goldring M.B., Cobelli N.J, & Sun H.B. (2016). Strain-induced mechanotransduction through primary cilia, extracellular ATP, purinergic calcium signaling, and ERK1/2 transactivates CITED2 and downregulates MMP-1 and MMP-13 gene expression in chondrocytes. Osteoarthritis and cartilage, 24(5).
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