Xenograft tumors of HER3/RH7777, RH7777, and CTOS C45 were excised and fixed in neutralized 10% formalin, embedded in paraffin, and cut into 4-μm-thick sections. The sections were incubated with diluted mouse anti-HER3 monoclonal antibody (2.5 μg/mL; erbB3/Her3, nanoTools, Teningen, Germany) for 1 h at room temperature, washed, and then incubated with peroxidase-conjugated secondary antibodies (EnVision+ System-HRP Labeled Polymer Anti-mouse; DAKO, Glostrup, Denmark). The binding of the secondary antibodies was visualized with diaminobenzidine staining (K3465; DAKO) and counterstaining was carried out with hematoxylin.
K3465
Manufactured by Agilent Technologies
The K3465 is a high-performance laboratory instrument designed for precision measurements. It offers advanced capabilities and functionalities for a variety of scientific and research applications. The core function of the K3465 is to provide accurate and reliable data collection and analysis.
Automatically generated - may contain errors
Lab products found in correlation
Envision system hrp labeled polymer anti mouse, by Agilent Technologies (1 mentions)
K0690, by Agilent Technologies (1 mentions)
S2020, by Agilent Technologies (1 mentions)
Mayer s hematoxylin, by Agilent Technologies (1 mentions)
S2o23 real, by Agilent Technologies (1 mentions)
S0809 antibody diluent, by Agilent Technologies (1 mentions)
3 protocols using k3465
1
Immunohistochemical Analysis of HER3 Expression
2
Immunolocalization of NeuGcGM3 Ganglioside
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Five-micrometer serial sections from each block were obtained in a micrometer (Leitz, 1512) and they were mounted on plus slides (Dako, S2024). All sections were attached to the slide by heating in a 60°C oven for 1 h. Afterward, the slides were dewaxed in xylene and rehydrated in graded ethanol series in the usual way. The samples were maintained in tap water until they were stained.
The immunolocalization of NeuGcGM3 ganglioside was performed as it was previously described in [10 (link)] with some modifications. Briefly, the slides were incubated with 14F7 Mab in a humid chamber for 1 h at room temperature followed by the labeled streptavidin biotin (LSAB) two steps' system (Dako, K0690) both for 30 minutes at room temperature. The enzymatic activity was visualized with 3,3-diaminobenzidine (DAB) substrate chromogenic solution (Dako, K3465) and the tissues were counterstained with Mayer's Hematoxylin (Dako, S2020). Concerning the evaluation of both EGFR and EGF tissue antigens, the procedure as it was previously described in [19 (link)] was used.
The immunolocalization of NeuGcGM3 ganglioside was performed as it was previously described in [10 (link)] with some modifications. Briefly, the slides were incubated with 14F7 Mab in a humid chamber for 1 h at room temperature followed by the labeled streptavidin biotin (LSAB) two steps' system (Dako, K0690) both for 30 minutes at room temperature. The enzymatic activity was visualized with 3,3-diaminobenzidine (DAB) substrate chromogenic solution (Dako, K3465) and the tissues were counterstained with Mayer's Hematoxylin (Dako, S2020). Concerning the evaluation of both EGFR and EGF tissue antigens, the procedure as it was previously described in [19 (link)] was used.
3
Histological Analysis of Keloid Biopsies
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Biopsy samples of keloids were collected and fixed with 4% paraformaldehyde and embedded in paraffin according to standard protocols. Hematoxylin/eosin staining and immunohistochemical staining (IHC) was performed from 6-µm thick paraffin sections as previously described (15, 16) . Briefly, Formalin fixed, paraffin-embedded tissue sections were deparaffinized, processed with appropriate antigen retrieval solution, incubated with the blocking reagent, and endogenous peroxidase activity was suppressed with hydrogen peroxide. Tissue sections were incubated with the primary antibody overnight at 4°C. The rat anti-CD31 (clone 550274) and rat anti-Ki67 (clone M7249 TEC-3) antibodies were used for IHC (both BD Pharmingen, Oxford, UK) followed by the appropriate horseradish peroxidase-conjugated (HRP) secondary antibodies (15) . The blocking reagents used for IHC were S2O23 REAL and S0809 Antibody Diluent (DakoCytomation). The peroxidase reactive chromogen used was 3.3 diaminobenzidine (DAB; K3465,DAKO, Agilent Technologies). Each staining experiment included sections stained without primary antibody as negative controls.
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