Total protein was extracted using radio immunoprecipitation assay (RIPA) lysis buffer with freshly added protease inhibitor (Roche, Basel, Switzerland). Western blot analysis was performed as described previously17 (link). Briefly, 40 μg total protein extract was loaded into a 10% sodium dodecyl sulphate polyacrylamide gel (SDS-PAGE) followed by transfer onto a polyvinylidene difluoride (PVDF) membranes (Roche, Basel, CH). Membranes were blocked with 5% skim milk, incubated first with rabbit polyclonal antibodies against human SET8 (1:1000; Abcam, Cambridge, UK), p53 (1:2,000; Santa Cruz, CA), anti-p53K382me1(1:1000; Affinity Biosciences, Cincinnati, OH)11 (link), or β-actin (1:10,000; Santa Cruz, CA) overnight at 4 °C, followed by incubation with secondary HRP-conjugated anti-rabbit IgG antibody (1:5000; Thermo Fisher, New York, NY). Proteins were visualized with an enhanced chemiluminescence reagent (Thermo Fisher, New York, NY) using the FluorChem® HD2 protein imprinting imaging system (Alpha InnoTec, San Leandro, CA).
Hrp conjugated anti rabbit igg antibody
Manufactured by Thermo Fisher Scientific
Sourced in United States
The HRP-conjugated anti-rabbit IgG antibody is a secondary antibody used in various immunoassay techniques. It is composed of an anti-rabbit IgG antibody conjugated with the enzyme horseradish peroxidase (HRP). This antibody can bind to and detect rabbit primary antibodies, allowing for the visualization and quantification of target proteins or antigens in a sample.
Automatically generated - may contain errors
Lab products found in correlation
Enhanced chemiluminescence reagent, by Thermo Fisher Scientific (2 mentions)
β actin, by Santa Cruz Biotechnology (2 mentions)
Protease inhibitor, by Roche (2 mentions)
Pvdf membrane, by Roche (1 mentions)
Total protein extraction kit, by Invent Biotechnologies (1 mentions)
Streptavidin peroxidase polymer, by Merck Group (1 mentions)
2019 ncov spike rbd antibody, by Sino Biological (1 mentions)
Nunc maxisorp, by Thermo Fisher Scientific (1 mentions)
Hrp conjugated anti mouse igg antibody, by Thermo Fisher Scientific (1 mentions)
G box, by Syngene (1 mentions)
Enhanced chemi luminescence (ecl), by Bio-Rad (1 mentions)
Ecl western blot kit, by Thermo Fisher Scientific (1 mentions)
5 protocols using hrp conjugated anti rabbit igg antibody
1
Western Blot Analysis of Protein Markers
2
Protein Extraction and Western Blot Analysis
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The protein of HCC tissues collected from High-copy HBV group and control group were extracted by Total Protein Extraction Kit (Invent Biotechnologies, USA) according to the manufacturer's instructions.
And the cell protein of CD8 + T cells collected from High-copy HBV group and control group were extracted by radio immunoprecipitation assay (RIPA) lysis buffer containing 1% protease inhibitor (Roche, Basel, Switzerland). Protein extract was subjected to 10% sodium dodecyl sulphate polyacrylamide gel (SDS-PAGE) followed by transfer onto a polyvinylidene di uoride (PVDF) membranes (Roche, Basel, CH). After blocking with 5% skim milk, the membrane was incubated with primary antibody against human PD-1/CD279 (PROTEINTECH, Chicago, USA) or β-actin (Santa Cruz, CA, USA) overnight at 4°C, followed by incubation with secondary HRP-conjugated anti-rabbit IgG antibody (Thermo Fisher, New York, USA). The relative intensities of protein bands were visualized with an enhanced chemiluminescence reagent (Thermo Fisher, New York, USA) using the FluorChem® HD2 protein imprinting imaging system (Alpha InnoTec, San Leandro, CA).
And the cell protein of CD8 + T cells collected from High-copy HBV group and control group were extracted by radio immunoprecipitation assay (RIPA) lysis buffer containing 1% protease inhibitor (Roche, Basel, Switzerland). Protein extract was subjected to 10% sodium dodecyl sulphate polyacrylamide gel (SDS-PAGE) followed by transfer onto a polyvinylidene di uoride (PVDF) membranes (Roche, Basel, CH). After blocking with 5% skim milk, the membrane was incubated with primary antibody against human PD-1/CD279 (PROTEINTECH, Chicago, USA) or β-actin (Santa Cruz, CA, USA) overnight at 4°C, followed by incubation with secondary HRP-conjugated anti-rabbit IgG antibody (Thermo Fisher, New York, USA). The relative intensities of protein bands were visualized with an enhanced chemiluminescence reagent (Thermo Fisher, New York, USA) using the FluorChem® HD2 protein imprinting imaging system (Alpha InnoTec, San Leandro, CA).
3
Quantifying SARS-CoV-2 Antibody and Chemokine Levels
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ELISA plates (Nunc MaxiSorp, Thermo Fisher) were coated with anti-IgG CH3 antibody (Bio-Rad, Hercules, CA; 1:500) overnight at 4 °C, followed by blocking with 4% bovine serum albumin (BSA) in PBS for 1 h at RT (room temperature). The supernatants were added to the wells and incubated for 1 h at 37 °C. For detection of CCL3L1, plates were washed and incubated with biotinylated anti-human MIP-1α (R&D Systems, Minneapolis, MN; 1:1,000), followed by streptavidin-peroxidase polymer (Sigma-Aldrich, St Louis, MO; 1:3,000). For detection of RBD, plates were incubated with rabbit anti-SARS-CoV-2 (2019-nCoV) Spike RBD Antibody (Sino Biological, Beijing, China; 1:1,000) followed by HRP-conjugated anti-rabbit IgG antibody (Invitrogen, Waltham, MA; 1:5,000). All incubations were performed for 1 h at 37 °C. The plates were then washed and TMB (3,3ʹ,5,5ʹ-tetramethylbenzidine) solution was added (Merck, Darmstadt, Germany). Color development was stopped after 10 min by adding 1 M HCl. Optical density was measured at 450 nm using a Tecan SPARK plate reader (Bergman Diagnostika AS, Kjeller, Norway).
4
Western Blot Analysis of Leishmania Proteins
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Leishmania cell lysates were prepared by re-suspending and boiling the cell pellets for 5 minutes in an SDS-PAGE sample buffer. Samples were resolved on SDS-polyacrylamide gel by electrophoresis and electro-blotted onto nitrocellulose membrane (0.45μm, Synergy scientific services) in Tris-glycine buffer (pH 8.3) at 80V for 3 hours. The membrane was treated with 5% skimmed milk to block the nonspecific sites, and then probed with rabbit anti-LdPfn antibodies [11 (link)] (1:100 dilution) or mouse anti-β tubulin monoclonal antibodies (Sigma, cat.No. T7816) (1:5000) or rabbit anti-eIF4A.1 antibodies (Cell Signalling Technology, cat No. 2490) (1:100) for overnight at 4°C. The membrane was washed 5 times with Tris-buffered saline (pH 7.5) containing 0.05% (v/v) Tween 20, then incubated with HRP conjugated anti-rabbit IgG antibody (Invitrogen, cat no. A16074) or HRP conjugated anti-mouse IgG antibody (Invitrogen, cat no.62-6820) for 2 hours and developed with ECL (Bio-Rad, Clarity) and imaging software (Syngene, G-box).
5
Western Blot Analysis of Purified Parasites
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