Immunohistochemistry was performed in human and murine colon sections in order to determine the NLRP3 expression level. The tissue samples were immersed in 4% paraformaldehyde for 24 h and then embedded in paraffin for immunohistochemistry analysis. The tissues were sectioned into slices and then were dried in a drying oven at 60°C for a half hour. Anti-NLRP3 antibody (GB11300; Servicebio, Wuhan, China) was detected using anti-rabbit secondary antibody for a half hour at room temperature. The tissue slices were counterstained using a DAB chromogenic reagent kit. Then the intensity of NLRP3 staining was analyzed using ImageJ Java.
Gb11300
Manufactured by Wuhan Servicebio Technology
Sourced in China
The GB11300 is a laboratory equipment designed for general use in research and diagnostic applications. It serves as a versatile tool for various scientific procedures. The core function of the GB11300 is to provide a controlled environment for conducting experiments or assays. Further details about the specific intended use or capabilities of this product are not available.
Automatically generated - may contain errors
Lab products found in correlation
Gb11383, by Wuhan Servicebio Technology (2 mentions)
Bm4813, by Boster Bio (1 mentions)
Ba2811, by Boster Bio (1 mentions)
Gb111364, by Wuhan Servicebio Technology (1 mentions)
Ab33985, by Abcam (1 mentions)
Gb11027, by Wuhan Servicebio Technology (1 mentions)
Fluorescence microscope, by Olympus (1 mentions)
Gb11131, by Wuhan Servicebio Technology (1 mentions)
G1012 100ml, by Wuhan Servicebio Technology (1 mentions)
Gb11142, by Wuhan Servicebio Technology (1 mentions)
Gb14073, by Wuhan Servicebio Technology (1 mentions)
Gb21301, by Wuhan Servicebio Technology (1 mentions)
Gb11022, by Wuhan Servicebio Technology (1 mentions)
Gb11119, by Wuhan Servicebio Technology (1 mentions)
Gb22303, by Wuhan Servicebio Technology (1 mentions)
Gb111841, by Wuhan Servicebio Technology (1 mentions)
Ab108334, by Abcam (1 mentions)
Gb11373, by Wuhan Servicebio Technology (1 mentions)
Triton x 100, by Solarbio (1 mentions)
Paraformaldehyde (pfa), by Wuhan Servicebio Technology (1 mentions)
7 protocols using gb11300
1
NLRP3 Expression in Colon Tissues
2
Renal Inflammation Pathway Analysis
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Renal paraffin sections were dewaxed and processed according to the standard experimental procedure, and the samples were incubated with the anti-NLRP3 (1 : 400, Servicebio, China, GB11300), anti-caspase-1 (1 : 1000, Servicebio, China, GB11383), anti-IL-1β (1 : 600, Servicebio, China, GB11113), anti-SOD2 (1 : 200, Boster, China, BM4813), and anti-NOX2 (1 : 600, Boster, China, BA2811) antibodies overnight at 4°C. The relative expression of the corresponding proteins was analyzed with ImageJ software (National Institutes of Health, USA).
3
Immunohistochemical Analysis of Renal Fibrosis
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Four-micron-thick renal paraffin sections were used for IHF staining. Briefly, after paraffin section dewaxing, rehydration, antigen repair, permeability, and blocking, the renal tissues were incubated with anti-F4/80 rabbit polyclonal antibody (Servicebio, GB11027, 1:500, Wuhan, China), anti-fibronectin (FN) rabbit polyclonal antibody (Servicebio, GB114057, 1:200, Wuhan, China), anti-α-SMA rabbit polyclonal antibody (Servicebio, GB111364, 1:400, Wuhan, China) and anti-NLRP3 rabbit polyclonal antibody (Servicebio, GB11300, 1:600, Wuhan, China) at 4°C overnight. After rewarming, the kidney tissue was incubated with a secondary antibody at room temperature for 1 hour. After staining the nucleus, the renal paraffin sections were observed and photographed under a fluorescence microscope. For costaining, after dewaxing, rehydration, antigen repair, permeability and blocking, anti-LC3B (Proteintech, 14600-1-AP, 1:200) antibody and anti-COXIV antibody (Abcam, ab33985, 1:500) were incubated in renal tissue simultaneously at 4°C overnight. After rewarming, the kidney tissue was incubated with anti-mouse and anti-rabbit secondary antibodies at room temperature for 1 hour simultaneously. Then the nuclei were stained and photographed.
4
Immunofluorescence Staining of Disc Cells
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Our previous study also described this method in detailly [2 (link)]. After intervertebral disc sections or NP cells were well prepared, 0.1% Triton X-100 was employed to permeate samples for 5 min. Then, samples were blocked with 3-5% BSA for 60 min at 37°C. Then, the samples were incubated using primary antibody against γH2AX (GB111841, 1 : 200, Servicebio, Wuhan, China), aggrecan (GB11373, 1 : 500, Servicebio, Wuhan, China), collagen type II (GB14073, 1 : 500, Servicebio, Wuhan, China), iNOS (GB11119, 1 : 1000, Servicebio, Wuhan, China), collagen type I (GB11022, 1 : 500, Servicebio, Wuhan, China), NLRP3 (GB11300, 1 : 1000, Servicebio, Wuhan, China), CD206 (#24595, 1 : 800, Cell Signaling Technology, Inc. USA), MMP3 (GB11131, 1 : 500, Servicebio, Wuhan, China), AGT (ab108334, 1 : 200, Abcam, USA), Nrf2 (340675, 1 : 500, Zenbio, Chengdu, China), and p65 (GB11142, 1 : 200, Servicebio, Wuhan, China) at 4°C overnight. At the second day, samples were treated with fluorescence secondary antibody (GB22303, GB21301, Servicebio, Wuhan, China) for 1 h in the dark room. The nuclei were staining with DAPI solution (G1012-100ML, Servicebio, Wuhan, China). The fluorescence was detected using fluorescence microscope (Olympus, Japan).
5
Histological Evaluation of Mammary Tissue
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Mammary gland tissues were excised immediately after euthanasia and fixed overnight in 4% paraformaldehyde (Servicebio, Wuhan, China). Tissues were paraffin-embedded and sectioned (5 μm), with sections dewaxed in xylene and rehydrated in an alcohol series, prior to staining with hematoxylin and eosin (HE; Beyotime Biotechnology Co. Ltd, Shanghai, China). Histological scoring was performed, without knowledge of treatment group, to grade tissue necrosis, dislodged epithelial cells, polymorphonuclear neutrophilic granulocyte inflammation, and lymphocytic infiltration, as described [35 (link)].
For immunohistochemistry (IHC), sections were deparaffinized and rehydrated. Sodium citrate buffer (pH = 6.0) was used for antigen retrieval. Sections were washed 3 times in PBS, incubated in PBS with 0.5% Triton X-100 (Solarbio Biotechnology Co. Ltd., Beijing, China) for 15 min and then incubated in block solution. Then sections were immunostained with primary antibodies NLRP3 (Servicebio #GB11300, 1:1000) or Keap1 (Servicebio #GB11847, 1:2000) at 4 °C overnight. Thereafter, sections were incubated with HRP-conjugated secondary antibodies (Servicebio #GB23303, 1:200) and stained with 3,3’-diaminobenzidine (DAB) (Servicebio #G1211).
For immunohistochemistry (IHC), sections were deparaffinized and rehydrated. Sodium citrate buffer (pH = 6.0) was used for antigen retrieval. Sections were washed 3 times in PBS, incubated in PBS with 0.5% Triton X-100 (Solarbio Biotechnology Co. Ltd., Beijing, China) for 15 min and then incubated in block solution. Then sections were immunostained with primary antibodies NLRP3 (Servicebio #GB11300, 1:1000) or Keap1 (Servicebio #GB11847, 1:2000) at 4 °C overnight. Thereafter, sections were incubated with HRP-conjugated secondary antibodies (Servicebio #GB23303, 1:200) and stained with 3,3’-diaminobenzidine (DAB) (Servicebio #G1211).
6
Immunohistochemical Analysis of Caspase-1 and NLRP3 in Cerebral Tissues
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Frozen sections were boiled in ethylenediaminetetraacetic acid (EDTA) antigen repair solution (pH 8.0) (Servicebio, Wuhan, CNH) for antigen retrieval and blocked with 3% bovine serum albumin for 30 min at room temperature. Then, the sections were incubated at 4°C overnight with primary antibodies against caspase-1 (1:50, Servicebio, Wuhan, CNH, GB11383) and NLRP3 (1:100, Servicebio, Wuhan, CNH, GB11300). Then, the sections were incubated with the corresponding secondary antibody at room temperature for 90 min. The sections were washed in PBS 3 times (5 min/time), and 4',6-diamidino-2-phenylindole (DAPI) (Servicebio, Wuhan, CNH, GB1012) was used for staining the dried section for 10 min. Apoptosis was detected by a fluorescein (FITC) TUNEL cell apoptosis detection kit (Servicebio, Wuhan, CNH, G1501-50T). Morphological changes in the cerebral tissues were observed under a microscope (Jiangnan Optical Instrument Group, Nanjing, China), and the images were collected for analysis (AlphaEaseFC). Two 90 × fields of view in the hippocampal CA1 area were randomly selected for photography. The number of positive cells was calculated according to the following formula: double-stained cells/total cells × 100%.
7
Immunohistochemical Analysis of NLRP3, Caspase-1, and GSDMD in Heart
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To determine the expression of NLRP3, caspase-1, and GSDMD in the heart, immunohistochemical analysis was performed using anti-NLRP3 (GB11300, Servicebio, Wuha, China), anti-caspase-1 (sc-56036, Santa Cruz, Dal-las, TX, USA), and anti-GSDMD (sc-81868, Santa Cruz, Dallas, TX, USA) antibodies as previously described [33] . Briefly, after dewaxing and antigen repair, the paraffin sections were placed in 3% hydrogen peroxide for 25 min at RT. The sections were subsequently incubated with primary antibodies overnight (16-18 h) at 4 °C and secondary antibodies at RT for 50 min. The sections were stained with diaminobenzidine (G1211, Servicebio, Wuha, China) and hematoxylin (G1004, Servicebio, Wuha, China). After sufficient drying, the slides were photographed and evaluated using the ImageJ software.
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