Proteins were extracted from the hippocampus of wild-type and heterozygous Tg2576 mice at postnatal days 180 and 360. Proteins were resolved by 4–12% sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Proteins were transferred (semi-dried: 40 minutes at 10 V; wet: overnight at 15 V) to polyvinylidene difluoride membranes. Non-specific binding was blocked with 5% skim milk powder in Tris-buffered saline containing 0.2% Tween 20. Membranes were incubated (overnight at 4°C) with the primary antibody, mouse anti-reelin monoclonal antibody (1:1,000; Chemicon, Billerica, MA, USA). After washing, membranes were then incubated with horseradish peroxidase-conjugated goat anti-mouse IgG (1:10,000; Millipore, Billerica, MA, USA) in conjunction with an enhanced chemiluminescence system (ECL Plus Western Blotting Detection System; GE Healthcare)[60 (link)]. Mouse anti-β-actin monoclonal antibody (Sigma-Aldrich) served as the internal control. The absorbance ratio of reelin-positive bands to β-actin represented the relative expression level of reelin protein.
Horseradish peroxidase conjugated goat anti mouse igg
Manufactured by Merck Group
Sourced in United States, Germany
Horseradish peroxidase-conjugated goat anti-mouse IgG is a secondary antibody reagent that consists of goat-derived polyclonal antibodies specific to mouse immunoglobulin G (IgG) molecules, which are conjugated to the enzyme horseradish peroxidase. This reagent can be used to detect and quantify the presence of mouse IgG in various immunoassay applications.
Automatically generated - may contain errors
Lab products found in correlation
Goat anti rabbit igg, by Merck Group (11 mentions)
1 2 dipalmitoyl sn glycero 3 phosphocholine, by NOF corporation (6 mentions)
N carbonyl methoxypolyethyleneglycol 2000 1 2 distearoyl sn glycero 3 phosphoethanolamine, by NOF corporation (6 mentions)
Protease inhibitor cocktail, by Merck Group (4 mentions)
Microplate reader, by Agilent Technologies (2 mentions)
3 3 5 5 tetramethylbenzidine substrate, by Thermo Fisher Scientific (2 mentions)
Maxisorp elisa plate, by Thermo Fisher Scientific (2 mentions)
1 2 dioleoyl 3 trimethylammonium propane, by Lipoid (2 mentions)
D type cpg odn, by Ajinomoto Group (2 mentions)
Nitrocellulose membrane, by GE Healthcare (2 mentions)
Ecl western blotting analysis system, by Merck Group (2 mentions)
Gapdh, by Cell Signaling Technology (2 mentions)
Biospot analyzer, by Cellular Technology (2 mentions)
Trueblue peroxidase substrate, by Cellular Technology (2 mentions)
Saponin, by Merck Group (2 mentions)
β actin, by Merck Group (2 mentions)
Pvdf membrane, by Merck Group (2 mentions)
Anti β actin antibody, by Merck Group (2 mentions)
Polyvinylidene difluoride membrane, by Merck Group (2 mentions)
Ne per nuclear and cytoplasmic extraction reagent, by Thermo Fisher Scientific (2 mentions)
51 protocols using horseradish peroxidase conjugated goat anti mouse igg
1
Reelin protein expression in Tg2576 mice
2
Protein Extraction and Western Blotting Protocol
3
Anti-RpfB Antibody Quantification by ELISA
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Serum was collected 4 weeks after the final immunization, and anti-RpfB antibody was assessed by indirect enzyme-linked immunosorbent assay (ELISA). A Maxisorp microtiter plate (Nunc, Roskilde, Denmark) was coated with 10 µg/mL of rRpfB protein and blocked with 5% fetal bovine serum (FBS)-0.05% Tween-phosphate buffered saline (PBST). Sera of the immunized mice were serially diluted and dispensed into each well. After 2 hours of incubation, horseradish peroxidase-conjugated goat anti-mouse IgG (Millipore, Bedford, MA, USA) was added. One hour later, color reaction was performed with 3,3', 5,5'-tetramethylbenzidine (Sigma) for 15 minutes. The reaction was stopped by 2 M H2SO4, and optical density was read at 450 nm.
4
SARS-CoV-2 Antibody Binding Assay
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Recombinant Wuhan-1, BA.4/5 receptor binding domain (RBD), or BA.4/5 spike protein (4 μg/mL) was immobilized on 96-well Maxisorp ELISA plates (Thermo Fisher) overnight at 4°C in coating buffer (1X PBS supplemented with 0.05% Tween-20, 2% BSA, and 0.02% NaN3). Plates were washed with PBS and blocked with 4% BSA for one hour at 25°C. Serum was serially diluted in 2% BSA and incubated on plates for 1 h at 25°C. After washing with PBS, RBD or spike-bound serum antibodies were detected with horseradish peroxidase conjugated goat anti-mouse IgG (1:500 dilution, Milipore Sigma) incubating for 2 h at 25°C. Plates were washed and developed with 3,3’−5,5’ tetramethylbenzidine substrate (Thermo Fisher), halted with 2 N H2SO4 and read at 450 nm using a microplate reader (BioTek). Mean serum endpoint titers were calculated with curve fit analysis of optical density (OD) values set as the reciprocal value of the serum dilution equal to the mean plus six times the standard deviation of background signal.
5
SARS-CoV-2 RBD and Spike Protein Antibody Binding Assay
6
Immunohistochemical Detection of Bcl-2 Protein
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Paraffin-embedded tissues were sectioned (4 µm thick) and mounted on glass slides. They were then dewaxed followed by antigen retrieval in 0.01M sodium citrate at 95°C for 10 min. Immunohistochemistry was done using mouse monoclonal anti–Bcl-2 (C-2) (IgG1; 1∶100; Santa Cruz Biotechnology) in blocking reagent at 25°C for 20 min. The primary antibody was omitted for negative controls. The slides were then treated consecutively with horseradish peroxidase–conjugated goat anti–mouse IgG (1∶200; Chemicon International, Inc., Temecula, CA, USA) and incubated for 1 h at room temperature. Slides were then incubated using a DAB substrate kit (Roche Applied Science, Indianapolis city, IN, USA), and the color reaction was allowed to develop for 5–10 min. After washing, slides were coverslipped.
7
Immunohistochemistry of Leptin Receptor and Glucose Transporters
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Lymphocytes (about 5 × 104 from each group) were dried and endogenic peroxidase was blocked by adding 200 μL of 3 % H2O2 solution. Then lymphocytes were placed in blocking buffer (1 % bovine serum albumin in PBS) with 2 % goat serum (Sigma). After 30 min mouse monoclonal antibody (1:200) against extracellular domain of human leptin receptor (R&D Systems) was added and then horseradish peroxidase-conjugated goat anti-mouse IgG (1:1000) (Chemicon International Inc. Ca). The negative control sample (for exclusion of non-specific binding of antibodies) consisted of lymphocytes incubated without the first antibody. The same procedure was used in the case of glucose transporters. The antibodies used were rabbit polyclonal antibody aimed against intracellular C-terminus of human GLUT1, GLUT3 and GLUT4 (1:100) and horseradish peroxidase-conjugated goat anti-rabbit IgG (1:2000) (Chemicon International Inc. Ca).
The antigen–antibody complex was visualized using DAB according to the manufacturers’ instructions (Sigma-Aldrich). The presence of investigated proteins was assessed using a light microscope (800×).
The antigen–antibody complex was visualized using DAB according to the manufacturers’ instructions (Sigma-Aldrich). The presence of investigated proteins was assessed using a light microscope (800×).
8
Influenza Vaccine Formulation Protocol
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1,2-dioleoyl-3-trimethylammonium-propane was purchased from Lipoid GmbH (Ludwigshafen, Germany). 1,2-dipalmitoyl-sn-glycero-3-phosphocholine and N-(carbonyl-methoxypolyethyleneglycol 2000)-1,2-distearoyl-sn-glycero-3-phosphoethanolamine were purchased from NOF Corporation (Tokyo, Japan). Cholesterol was purchased from Fujifilm Wako Pure Chemical Corporation, Ltd. (Osaka, Japan). D-type CpG ODN (CpG ODN: 5′-ggtgcatcgatgcagggggg-3′) was purchased from GeneDesign (Osaka, Japan). Horseradish-peroxidase-conjugated goat anti-mouse IgG was purchased from Merck Millipore (Darmstadt, Germany). Horseradish-peroxidase-conjugated goat anti-mouse IgG1, IgG2b, and IgG2c were purchased from SouthernBiotech (Birmingham, AL, USA). Alhydrogel adjuvant 2%, as alum, was purchased from InvivoGen (San Diego, CA, USA). Ether-treated, HA-antigen-enriched, virion-free SV from H1N1 influenza A virus (strain: A/California/7/2009 (Cal7)) was kindly provided by Dr. Yasuyuki Gomi of the Research Foundation for Microbial Diseases of Osaka University, Osaka, Japan. The SV contained 570 μg of HA in 1000 μg of SV. H1N1 influenza A virus (strain: Cal7) was kindly provided by Dr. Hideki Asanuma of the National Institute of Infectious Diseases, Tokyo, Japan.
9
H1N1 Influenza Vaccine Protocol
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Horseradish-peroxidase–conjugated goat anti-mouse IgG was purchased from Merck Millipore (Darmstadt, Germany). Alhydrogel adjuvant (2%) was purchased from InvivoGen (San Diego, CA, USA). K3 CpG oligodeoxynucleotide (ODN; 5′-atcgactctcgagcgttctc-3′) was purchased from Gene Design (Osaka, Japan). H1N1 influenza A virus (strain: A/California/7/2009 [Cal7]) was kindly provided by Dr. Hideki Asanuma (National Institute of Infectious Diseases, Tokyo, Japan).
10
Western Blot Analysis of Bax Protein in Neurons
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Protein level of Bax in neurons was examined by western blot analysis according to the product protocol. Briefly, neurons of each group were harvested and lysed in radioimmunoprecipitation assay (RIPA) buffer (Beyotime Inc., Shanghai, China) which contained protease inhibitors to obtain the total protein from the cells. Protein concentration was determined using the bicinchoninic acid protein assay kit (Beyotime Inc.). 40 μg of protein from the cell lysate in each group was run per lane on 12% sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) gels, electrophoresed, and transferred to a nitrocellulose filter membrane. After the membranes were blocked with 5% nonfat dry milk, they were incubated overnight at 4°C with rabbit anti-β-actin (1:1,000; Abcam, Hong Kong, China) and rabbit anti-Bax (1:1,000; Abcam). Horseradish peroxidase-conjugated goat anti-mouse IgG was used as secondary antibody (1:2,500; Merck, Darmstadt Germany). The membranes were incubated for 2 hours at 25°C. The immunoreactive bands were detected by the SuperSignal West Pico chemiluminescence detection system (Pierce, Rockford, IL, USA). Densitometric analysis was performed by NIH Image J analysis software (National Institutes of Health, Bethesda, MD, USA). The values were normalized with β-actin.
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