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Horseradish peroxidase conjugated donkey anti mouse igg

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Horseradish peroxidase-conjugated donkey anti-mouse IgG is a secondary antibody conjugated with the enzyme horseradish peroxidase. It is designed to bind to mouse IgG antibodies and can be used in various immunoassay techniques, such as Western blotting, ELISA, and immunohistochemistry.

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Lab products found in correlation

Sureblue tmb, by LGC (2 mentions) Horseradish peroxidase conjugated donkey anti goat igg, by Jackson ImmunoResearch (1 mentions) Cy3 conjugated donkey anti rabbit igg, by Jackson ImmunoResearch (1 mentions) Horseradish peroxidase conjugated donkey anti rabbit igg, by Jackson ImmunoResearch (1 mentions) Dextran sulfate sodium (dss), by MP Biomedicals (1 mentions) Opteia assay diluent, by BD (1 mentions) Chrompure, by Jackson ImmunoResearch (1 mentions) Alexa fluor 488 conjugated ib4, by Thermo Fisher Scientific (1 mentions) Alexa fluor 488 conjugated goat anti rat igg, by Thermo Fisher Scientific (1 mentions) Alexa fluor 594 conjugated goat anti rat igg, by Thermo Fisher Scientific (1 mentions) Alexa fluor 594 conjugated donkey anti rabbit igg, by Thermo Fisher Scientific (1 mentions) Trueblue peroxidase substrate, by LGC (1 mentions) S6 universal analyzer, by Cellular Technology (1 mentions) Immunospot software, by Cellular Technology (1 mentions) Methylcellulose, by Merck Group (1 mentions) Tissuelyser 2, by Qiagen (1 mentions)

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Horseradish peroxidase

7 protocols using horseradish peroxidase conjugated donkey anti mouse igg

1

Quantifying Type I Collagen Production

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Type 1 collagen production was determined by western blotting. Monolayers of MC3T3-E1 cells were lysed directly in 1X Laemmli sample buffer, dispersed by sonication, and resolved by sodium dodecyl sulfate—polyacrylamide gel electrophoresis. Immune complexes on nitrocellulose membranes were detected using mouse monoclonal anti-type I collagen IgG1 (COL1A: sc59772; Santa Cruz Biotechnology, Santa Cruz, CA) with horseradish peroxidase-conjugated donkey anti-mouse IgG (Code: 715-035-150; Jackson ImmunoResearch Laboratories Inc., West Grove, PA) as secondary antibody. The cell content of each sample was determined by western blotting in parallel for β-actin using mouse monoclonal anti-actin IgG1 (C-2: sc8432; Santa Cruz Biotechnology, Santa Cruz, CA). Immune complexes were visualized on X-ray film by enzyme-linked chemiluminescence and quantified using a Fluor-S MultiImager. Data were expressed as the ratio of type I collagen to actin in each sample.
Luttrell L.M., Dar M.S., Gesty-Palmer D., El-Shewy H.M., Robinson K.M., Haycraft C.J, & Barth J.L. (2019). Transcriptomic characterization of signaling pathways associated with osteoblastic differentiation of MC-3T3E1 cells. PLoS ONE, 14(1), e0204197.
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2

Antibody Reagents for Western Blotting

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The primary antibodies used in the studies are listed in Supplementary Table 2. Horseradish-peroxidase–conjugated donkey anti-mouse IgG, horseradish-peroxidase–conjugated donkey anti-rabbit IgG, horseradish-peroxidase–conjugated donkey anti-goat IgG, and Cy3-conjugated donkey anti-rabbit IgG were from Jackson Immuno Research (West Grove, PA). DSS (molecular weight, 36–50 kilodaltons) was purchased from MP Biomedicals (Solon, OH).
Watanabe N., Mashima H., Miura K., Goto T., Yoshida M., Goto A, & Ohnishi H. (2016). Requirement of Gαq/Gα11 Signaling in the Preservation of Mouse Intestinal Epithelial Homeostasis. Cellular and Molecular Gastroenterology and Hepatology, 2(6), 767-782.e6.
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3

Quantification of Serum IgG and Viral Antibodies

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To measure total IgG levels and antibody-specificity in the serum, plates were coated with either 2 µg/ml of donkey anti-mouse IgG (Affinipure; Jackson ImmunoResearch Laboratories, West Grove, PA), carbonate buffer (0.0875 M Na2CO3, 0.0125 M HCO3, pH 9.2) or 0.5% paraformaldehyde-fixed viral antigen–carbonate buffer and incubated overnight at 4°C. Coated plates were washed in PBS with 0.05% Tween 20 (PBS-T), and blocked in 3% milk–PBS-T prior to incubation with serial dilutions of serum or the mouse IgG standard (Invitrogen, Jackson ImmunoResearch) in 1% milk-PBS-T for 2 hrs at RT. IgG was detected by the use of horseradish peroxidase-conjugated donkey anti-mouse IgG (Jackson ImmunoResearch), SureBlue TMB (SeraCare, Milford, PA), and stop solution, and absorbance at 450 nm was read on FluoStar Omega (BMT Labtech, Cary, NC).
Hogan C.H., Owens S.M., Reynoso G.V., Kirillov V., Meyer T.J., Zelazowska M.A., Liu B., Li X., Chikhalya A., Dong Q., Khairallah C., Reich N.C., Sheridan B., McBride K.M., Hearing P., Hickman H.D., Forrest J.C, & Krug L.T. (2023). B cell-intrinsic STAT3-mediated support of latency and interferon suppression during murine gammaherpesvirus 68 infection revealed through an in vivo competition model. bioRxiv.
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4

Quantifying Serum IgG Levels

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To measure total IgG levels in the serum, plates were coated with 2 µg/ml of donkey anti-mouse IgG (Affinipure; Jackson ImmunoResearch Laboratories, West Grove, PA)–PBS, washed in PBS with 0.05% Tween 20, and blocked in 1% bovine serum albumin (BSA)–PBS prior to incubation with serial dilutions of serum or the mouse IgG standard (ChromPure; Jackson ImmunoResearch) in assay diluent (OptEIA assay diluent; BD Biosciences) overnight at 4°C. IgG was detected by the use of horseradish peroxidase-conjugated donkey anti-mouse IgG (Jackson ImmunoResearch), R & D Systems (Minneapolis, MN) substrate, and stop solution, and absorbance at 450 nm was read on a FilterMax5 microplate reader (Molecular Devices, Sunnyvale, CA). To measure levels of IgG specific for viral antigens, plates were first coated with 0.5% paraformaldehyde-fixed viral antigen–PBS overnight at 4°C.
Reddy S.S., Foreman H.C., Sioux T.O., Park G.H., Poli V., Reich N.C, & Krug L.T. (2016). Ablation of STAT3 in the B Cell Compartment Restricts Gammaherpesvirus Latency In Vivo. mBio, 7(4), e00723-16.
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5

Quantifying Antibody Levels and Specificity

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To measure total IgG levels and antibody specificity in the serum, plates were coated with either 2 µg/mL of donkey anti-mouse IgG (Affinipure; Jackson ImmunoResearch Laboratories, West Grove, PA), carbonate buffer (0.0875 M Na2CO3, 0.0125 M HCO3, pH 9.2) or 0.5% paraformaldehyde-fixed viral antigen-carbonate buffer and incubated overnight at 4°C. Coated plates were washed in PBS with 0.05% Tween 20 (PBS-T) and blocked in 3% milk-PBS-T prior to incubation with serial dilutions of serum or the mouse IgG standard (Invitrogen, Jackson ImmunoResearch) in 1% milk-PBS-T for 2 h at RT. IgG was detected by the use of horseradish peroxidase-conjugated donkey anti-mouse IgG (Jackson ImmunoResearch), SureBlue TMB (SeraCare, Milford, PA), and stop solution, and absorbance at 450 nm was read on FluoStar Omega (BMT Labtech, Cary, NC).
Hogan C.H., Owens S.M., Reynoso G.V., Liao Y., Meyer T.J., Zelazowska M.A., Liu B., Li X., Grosskopf A.K., Khairallah C., Kirillov V., Reich N.C., Sheridan B.S., McBride K.M., Gewurz B.E., Hickman H.D., Forrest J.C, & Krug L.T. (2024). Multifaceted roles for STAT3 in gammaherpesvirus latency revealed through in vivo B cell knockout models. mBio, 15(2), e02998-23.
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6

Extracellular Matrix Protein Immunodetection

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The primary antibodies used in this study were as follows: anti-penta-His mouse monoclonal antibody (Qiagen), anti-mammalian type IV collagen rabbit polyclonal antibody (Rockland Immunochemicals, Pottstown, PA), anti-heparan sulfate proteoglycan (perlecan) rat monoclonal antibody (A7L6; Millipore), anti-EHS laminin rabbit polyclonal antibody (L9393; Sigma-Aldrich), and anti-mouse nidogen-1 rat monoclonal antibody (ELM1; Santa Cruz Biotechnology, Dallas, Tx). Horseradish peroxidase-conjugated donkey anti-mouse IgG (Jackson ImmunoResearch) was used as a secondary antibody for western blotting. Alexa Fluor™ 488-conjugated goat anti-rat IgG, Alexa Fluor™ 594-conjugated goat anti-rat IgG, Alexa Fluor™ 594-conjugated donkey anti-rabbit IgG, and Alexa Fluor™ 680-conjugated donkey anti-rabbit IgG (Thermo Fisher Scientific) were used for immunofluorescence histochemistry and Alexa Fluor ™ 488-conjugated IB4 (Thermo Fisher Scientific) for visualization of endothelial cells. DAPI or Hoechst33342 was used for nuclear counterstaining.
Futaki S., Horimoto A., Shimono C., Norioka N., Taniguchi Y., Hamaoka H., Kaneko M., Shigeta M., Abe T., Sekiguchi K, & Kondo Y. (2023). Visualization of basement membranes by a nidogen-based fluorescent reporter in mice. Matrix Biology Plus, 18, 100133.
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7

CCHFV Antigen Detection Assay

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Mouse tissues were dissected into tubes containing a steel bead and SW13 growth media and homogenized at 30hz using a TissueLyser II (Qiagen). Clarified tissue homogenates were diluted in L-15 media and then applied to SW13 cells in a 96-well format. Sample was then removed, and cells overlaid with L-15 media supplemented with 0.8% methylcellulose (Sigma). Cells were incubated at 37 °C for 40 h. Overlay was removed, and cells fixed with 2% paraformaldehyde overnight at 4 °C. To detect CCHFV antigen, mouse anti-CCHFV Gc monoclonal 11E7 (BEI Resources) was applied at 500 ng/mL. Plates were washed, and bound antibody detected with horseradish peroxidase conjugated donkey anti-mouse IgG (Jackson Immuno Research). Plates were developed with TrueBlue peroxidase substrate (Seracare) and foci counted with an S6 Universal analyzer and Immunospot software (Cellular Technology Limited).
Hawman D.W., Haddock E., Meade-White K., Williamson B., Hanley P.W., Rosenke K., Komeno T., Furuta Y., Gowen B.B, & Feldmann H. (2018). Favipiravir (T-705) but not ribavirin is effective against two distinct strains of Crimean-Congo hemorrhagic fever virus in mice. Antiviral research, 157.
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