We analyzed the RG, pull-down and hemolymph samples with standard Western blotting. The following antibodies were used to detect proteins on the blot. Flag-tagged proteins were detected with a monoclonal mouse anti-Flag antibody (Cell Signaling, #8146 S, 1:1000 dilution) followed by incubation with goat anti-mouse IgG H&L HRP secondary antibody (Abcam, #97023) at a ratio of 1:15,000. To detect Myc-tagged proteins, we used a monoclonal rabbit anti-Myc antibody (Cell Signaling, #2278 S, 1:1000 dilution), and blots were incubated with a goat anti-rabbit IgG H&L HRP secondary antibody (Abcam, #97051, 1:15,000 dilution). All protein bands were scanned and detected with the Bio-Rad ChemidocTM MP imaging system at the University of Alberta.
Goat anti mouse igg h l hrp secondary antibody
Manufactured by Abcam
Sourced in United Kingdom
Goat anti-mouse IgG H&L HRP secondary antibody is a polyclonal antibody that recognizes the heavy and light chains of mouse immunoglobulin G (IgG). The antibody is conjugated with horseradish peroxidase (HRP) enzyme, which can be used for detection in various immunoassays.
Automatically generated - may contain errors
Lab products found in correlation
Fetal bovine serum (fbs), by Thermo Fisher Scientific (2 mentions)
Chemidoc mp imaging system, by Bio-Rad (1 mentions)
Goat anti rabbit igg h l hrp secondary antibody, by Abcam (1 mentions)
Anti flag antibody, by Cell Signaling Technology (1 mentions)
Hiv type 1 p24 antigen elisa 2, by ZeptoMetrix (1 mentions)
Recombinant hiv 1 p24 protein, by Abcam (1 mentions)
Varioskan lux, by Thermo Fisher Scientific (1 mentions)
Sodium pyruvate, by Thermo Fisher Scientific (1 mentions)
Trypsin ethylenediaminetetraacetic acid, by Thermo Fisher Scientific (1 mentions)
Human hepg2 cells, by American Type Culture Collection (1 mentions)
Antibiotic antimycotic, by Thermo Fisher Scientific (1 mentions)
Caffeine, by Merck Group (1 mentions)
Polyvinylidene fluoride pvdf membrane, by Bio-Rad (1 mentions)
Adenosine triphosphate (atp), by Thermo Fisher Scientific (1 mentions)
Supersignal west dura chemiluminescent substrate, by Thermo Fisher Scientific (1 mentions)
2 4 2 hydroxyethyl piperazin 1 yl ethanesulfonic acid hepes, by Merck Group (1 mentions)
4 benzoylbenzoic acid, by Merck Group (1 mentions)
N methylmorpholine, by Merck Group (1 mentions)
4 protocols using goat anti mouse igg h l hrp secondary antibody
1
Western Blot Protein Detection
2
HIV-1 Gag p24 Quantification and Antibody Detection
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For HIV-1 Gag p24 determination, two days after transfection, p24 antigen in cell culture media was detected using HIV Type 1 p24 Antigen ELISA 2.0 (ZeptoMetrix) following kit instructions.
For plasma antibody determination, recombinant HIV-1 p24 protein (Abcam, 0.5 μg/ml) was coated in 96-well EIA/RIA plates overnight at 4 °C. The plates were blocked with 5% skim milk at 37 °C for 1 h. After washing, plasmas (two-fold serially diluted from 1:50 or 50-fold diluted) were added and incubated for 1 h at 37 oC. After washing, goat anti-mouse IgG H&L (HRP) secondary antibody (Abcam) diluted 1:50,000 was added. Plates were then incubated at 37 oC for 1 h. After extensive washes, 100 µl of the substrate was added and incubated for 10 min at room temperature, followed by adding 100 µl of stop solution. The optical density at 450 nm was determined using a Varioskan LUX (Thermo Scientific).
For plasma antibody determination, recombinant HIV-1 p24 protein (Abcam, 0.5 μg/ml) was coated in 96-well EIA/RIA plates overnight at 4 °C. The plates were blocked with 5% skim milk at 37 °C for 1 h. After washing, plasmas (two-fold serially diluted from 1:50 or 50-fold diluted) were added and incubated for 1 h at 37 oC. After washing, goat anti-mouse IgG H&L (HRP) secondary antibody (Abcam) diluted 1:50,000 was added. Plates were then incubated at 37 oC for 1 h. After extensive washes, 100 µl of the substrate was added and incubated for 10 min at room temperature, followed by adding 100 µl of stop solution. The optical density at 450 nm was determined using a Varioskan LUX (Thermo Scientific).
3
Effects of Caffeine on Hepatocyte Antioxidant Enzymes
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Human HepG2 cells were purchased from ATCC (Middlesex, UK). Caffeine was purchased from Sigma-Aldrich Co. (St. Louis, MO, USA). Minimum essential medium (MEM) with glutamine, fetal bovine serum, antibiotic-antimycotic, sodium pyruvate, trypsin-ethylenediaminetetraacetic acid and horseradish peroxidase (HRP) chemiluminescent substrate were purchased from Thermo Fisher (Waltham, MA USA). PON1, PON3, alpha tubulin primary antibodies and goat anti-mouse IgG H&L HRP secondary antibody were purchased from Abcam (Cambridge, UK). ApoA1 primary antibody was purchased from Novus Biologicals Inc. (Littleton, CO, USA). The radioimmunoprecipitation assay (RIPA) lysis buffer system was purchased from Santa Cruz (Heidelberg, Germany). The polyvinylidene fluoride (PVDF) membrane was purchased from Bio-Rad (Hercules, CA, USA). Other chemicals were purchased from Sigma-Aldrich Co. (St. Louis, MO, USA) or Merck (Darmstadt, Germany). All reagents were of analytical grade.
4
In Vitro CK2 Kinase Assay
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