The following main reagents were used: RNA Extraction kit (Xinmai Biotechnology Co., Ltd., Shanghai, China), RT-qPCR kit (Applied Biosystems, Foster City, CA, USA), rabbit anti-human RECK, P53, and RUNX monoclonal primary antibody (Acris Antibodies Inc., San Diego, CA, USA), mouse anti-rabbit polyclonal secondary antibody (HRP-labeled) (Genewiz, Suzhou, China), primary antibody and secondary antibody of GAPDH were purchased from Thermo Fisher Scientific (Waltham, MA, USA), immunohistochemistry kit (Roche, Indianapolis, IN, USA), ELISA kit (Takara, Dalian, China), methylation determination kit (Kang Century Biotech Co., Ltd., Beijing, China) and other chemical reagents were from Sinopharm Chemical Reagent Co., Ltd. (Shanghai, China). In addition, the following main instruments were used: Fluorescence quantitative PCR instrument (Applied Biosystems), microplate reader (Beijing Liuyi Biotechnology, Beijing, China), protein electrophoresis (Beijing Liuyi Biotechnology), gel imager (Bio-Rad, Hercules, CA, USA), Olympus microscope X53, Mindrop micro-nucleic acid quantitative instrument (Bio-Rad).
Immunohistochemistry kit
Manufactured by Roche
Sourced in United States
The Immunohistochemistry (IHC) kit is a laboratory equipment used for the detection and visualization of specific proteins or antigens in tissue sections. The kit provides reagents and protocols for the immunohistochemical staining process, enabling the identification and localization of target proteins within the tissue sample.
Automatically generated - may contain errors
Lab products found in correlation
Optical microscope, by Olympus (2 mentions)
Gel imager, by Bio-Rad (1 mentions)
Rt qpcr kit, by Thermo Fisher Scientific (1 mentions)
Fluorescence quantitative pcr instrument, by Thermo Fisher Scientific (1 mentions)
Lc3b ab, by Cell Signaling Technology (1 mentions)
Cd34 monoclonal antibody, by Abcam (1 mentions)
Biotinylated secondary antibody, by Abcam (1 mentions)
5 protocols using immunohistochemistry kit
1
Comprehensive Analysis of Molecular Biomarkers
2
Immunohistochemical Analysis of LC3B
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Heart sections from the Wistar rats were stained through the use of an immunohistochemistry kit (no.760-700, Roche Life Science, Indianapolis, USA), and the target protein was identified by brown color. As primary antibodies, LC3B Ab (#2775, Cell Signaling Technology, Maryland, USA) were used.
3
Immunohistochemical Quantification of Microvessel Density in Hepatocellular Carcinoma
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The microvessel density (MVD) was detected through immunohistochemistry (the immunohistochemistry kit was purchased from Hoffmann-La Roche Ltd) using CD34 monoclonal antibody. The slices of HCC tissues and adjacent nontumor tissues were taken for routine dewaxing, antigen retrieval, and sealing. After adding CD34 monoclonal antibody (dilution at 1:100; Abcam Co.) of 100 μL into each slice, the tissues were incubated at 4°C overnight. Then, the slices were washed with PBST and added with the biotin-labelled secondary antibody (dilution at 1:500; Abcam Co.) for a 2-hour incubation at 37°C. The streptavidin avidin biotin complex was added, and the slices were incubated at 37°C for 1 h. After DAB color reaction, hematoxylin was applied for a 2-min counterstaining. And the slices were sealed with neutral resin and observed under optical microscope (Olympus) in a low power (×100) for selection of areas with most new vessels. Cell counting was performed in a high power (×400). Each group was assigned with 6 fields, the average of which was calculated as the MVD. The endothelial cells or cell clusters that presented brown color in immunohistochemistry staining and displayed a distinct border with adjacent microvessels and tumor cells in tumor tissues were all new tumor vessels.
4
Immunohistochemical Analysis of ENC1 and TCF4
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The expression of ENC1 or TCF4 in paraffin-embedded tumor samples was checked using an immunohistochemistry kit (Roche) according to the manufacturer’s instructions. Briefly, tumor slides were treated with pepsin for 10 min and incubated with digoxin (DIG)-labeled monoclonal antibody (RiboBio, Guangzhou, Guangdong, China) for 4 h (both at ambient temperature). After three washes (5 min) with PBST and sealing with 10% FBS for 30 min, the sections were incubated overnight at 4 °C with anti-DIG antibody. Subsequently, the sections were treated with HRP-conjugated anti-rabbit IgG for 1 h. After the addition of diaminobenzidine and hematoxylin, the sections were finally observed under an optical microscope. The expression of ENC1 or TCF4 was analyzed by combining the percentage of positively stained tumor cells with the intensity of positive staining. Staining intensity was graded as follows: 0, no staining; 1, weak staining (light purple); 2, moderate staining (purple-dark purple); 3, strong staining (dark purple) as described by Cerilli et al. [23 (link)].
5
Immunohistochemical Analysis of HGF and c-Met in HCC
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The HCC tissues and adjacent nontumor tissues were obtained and fixed with 4% formalin. After gradient alcohol dehydration, the tissues were embedded with paraffin and then sliced to serial tissue sections of 5-μm thick. After routine dewaxing and antigen reparation, the tissues were sealed with blocking buffer. Then the slices were added with HGF or c-Met antibody (dilution at 1:500, 100 μL/slice; Abcam Co., Cambridge, MA, USA) and incubated at 4°C overnight. After washing by phosphate buffer solution Tween-20 (PBST), the biotinylated secondary antibody (dilution at 1:500; Abcam Co.) was added and the tissues were incubated at 37°C for 2 h. Avidin-Biotin-Enzyme Complex (ABC) were added for incubation at 37°C for 1 h. After diaminobenzidine (DAB) color reaction, the slices were blocked, observed, and photographed under optical microscope (Olympus, Southborough, MA, USA). The immunohistochemistry kit was purchased from Roche (Hoffmann-La Roche Ltd, Mannheim, Germany). Under the same light intensity, the HGF and c-Met in HCC tissues and adjacent nontumor tissues were examined, with a random selection of 4 fields assigned to each slice. The percentage of positive cells displayed in the immunohistochemical staining under the high power lens of ×400 was counted as an index. The positive rate = the number of positive cells/the number of cells in the fields ×100%.
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