Hrp conjugated anti mouse igg

Manufactured by Santa Cruz Biotechnology

Sourced in United States

HRP-conjugated anti-mouse IgG is a secondary antibody conjugated with horseradish peroxidase (HRP). It is designed to detect and bind to mouse immunoglobulin G (IgG) in various immunoassays and immunochemical techniques.

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Lab products found in correlation

Hrp conjugated anti rabbit igg, by Santa Cruz Biotechnology (9 mentions) Pvdf membrane, by Merck Group (7 mentions) Ripa buffer, by Thermo Fisher Scientific (4 mentions) Cleaved caspase 3, by Cell Signaling Technology (3 mentions) Nitrocellulose membrane, by GE Healthcare (3 mentions) Mes buffer, by Thermo Fisher Scientific (3 mentions) Anti β actin, by Merck Group (3 mentions) Anti erk, by Cell Signaling Technology (3 mentions) Hepg2 human hepatoma cells, by American Type Culture Collection (2 mentions) Halt protease and phosphatase inhibitor cocktail 100x, by Thermo Fisher Scientific (2 mentions) Nupage 4 12 bis tris gel in, by Thermo Fisher Scientific (2 mentions) Anti fosl2, by Cell Signaling Technology (2 mentions) Gradient mini protean tgx precast protein gels, by Bio-Rad (2 mentions) Anti fosl1, by Cell Signaling Technology (2 mentions) Trans blot turbo transfer pack, by Bio-Rad (2 mentions) Dc protein assay, by Bio-Rad (2 mentions) Bioruptor ucd 200, by Diagenode (2 mentions) Gapdh, by Santa Cruz Biotechnology (2 mentions) Horseradish peroxidase hrp conjugated anti rabbit igg, by Santa Cruz Biotechnology (2 mentions) Bca protein assay kit, by Abcam (2 mentions)

54 protocols using hrp conjugated anti mouse igg

Piceatannol Modulates Hepatic Signaling

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Piceatannol (purity > 98.0%) was purchased from Tokyo Chemical Industry (Portland, OR). HepG2 human hepatoma cells were obtained from American Type Culture Collection (Manassas, VA). Rabbit antibodies for acetyl-CoA carboxylase (ACC), phosphorylated ACC, extracellular signal-regulated kinases 1/2 (ERK1/2), phosphorylated ERK1/2, c-Jun N-terminal kinase (JNK), phosphorylated JNK and peroxisome proliferator-activated receptor alpha (PPARα) were purchased from Cell Signaling Technology (Danvers, MA). Mouse antibodies for peroxisome proliferator-activated receptor gamma (PPARγ), and secondary antibodies of horseradish peroxidase (HRP)-conjugated anti-rabbit IgG and HRP-conjugated anti-mouse IgG were obtained from Santa Cruz Biotechnology (Dallas, TX). Fetal bovine serum and fatty acid-free bovine serum albumin (BSA) were purchased from Sigma-Aldrich (St. Louis, MO). Penicillin/streptomycin was purchased from GE Healthcare (Marlborough, MA). All other chemicals were obtained with either analytical grade or cell-culture grade from Fisher Scientific (Waltham, MA).

Yang J.S., Tongson J., Kim K.H, & Park Y. (2020). Piceatannol attenuates fat accumulation and oxidative stress in steatosis-induced HepG2 cells. Current Research in Food Science, 3, 92-99.

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Extraction and Immunoblotting of Proteins from FFPE Tissue

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Deparaffinization and protein extraction of FFPE tissue was carried out using Qproteome FFPE Tissue Kit (Qiagen, Hilden, Germany; #1042481) following manufacturer’s instructions. Extracted proteins were determined by Bradford assay (BIO-RAD, CA, USA; #5000205). Quantified proteins were mixed with 5 × SDS-PAGE loading buffer (Biosesang, Seongnam, Korea; S2002) in concentration of 2 μg/μL and boiled at 95 °C for 5 min. Equal quantities of protein were separated to SDS-PAGE gel and transferred to nitrocellulose membranes (BIO-RAD; #1704158). Membranes were blocked by incubation in 5% skim milk in Tris-buffered saline (TBS) with 0.1% Tween-20 and probed with antibody against ATX (1:1000; Abcam, Cambridge, UK; ab140915), LPA1(1:2000; Abcam; ab166903), and LPA2(1:1000; Abcam; ab38322) diluted in 1% BSA in TBS, 0.1% Tween 20, and 0.02% NaN₃. The membranes were washed and then incubated with secondary antibodies (HRP conjugated anti-mouse IgG, or anti-rabbit IgG) (1:20,000; Santa Cruz, TX, USA) for 1 h at room temperature. The bands were visualized using WesternBright ECL (Advansata, CA, USA; K-12045-D50) after washing the membrane and exposed to X-ray film.

Cha Y.J, & Koo J.S. (2019). Expression of Autotaxin–Lysophosphatidate Signaling-Related Proteins in Breast Cancer with Adipose Stroma. International Journal of Molecular Sciences, 20(9), 2102.

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Western Blot Analysis of SOCS Proteins

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Adherent cells were lysed in cell lysis buffer, and Western blot analysis was performed as previously described (8 (link)). Cells were treated with the proteasome inhibitor MG132 (N-carbobenzyloxy-L-leucyl-L-leucyl-L-leucinal) (5 μM) for 16 hours before the lysis of cells for the analysis of SOCS4 and SOCS5 proteins. Antibodies used for Western blotting were as follows: anti-pSTAT3 (Y705), total STAT3, pERK (p44/42, T202/Y204), pAKT (S473), total AKT, pS6 (S240/244), S6, pEGFR (Y1068), total EGFR, L858R-EGFR– or del19-EGFR–specific antibodies, anti-JAK2 (Cell Signaling Technology), and anti-JAK1 (BD Biosciences); anti–total ERK, EGFR, Myc, ubiquitin and horseradish peroxidase (HRP)–conjugated anti-rabbit IgG, HRP-conjugated anti-mouse IgG, HRP-conjugated anti-rabbit IgG, SOCS4, and SOCS5 (Santa Cruz Biotechnology); anti–c-MET (Invitrogen); and anti–α-tubulin (Sigma-Aldrich). Densitometric quantification of high-resolution immunoblot images was performed using an analysis program, ImageJ (developed by W. Rasband, Research Services Branch of the National Institute of Mental Health). Analyses of the phosphorylation status of RTK and ErbB family members were performed using the RayBio Human RTK and EGFR Phosphorylation Array kits according to the manufacturer’s instruction (RayBiotech).

Gao S.P., Chang Q., Mao N., Daly L.A., Vogel R., Chan T., Liu S.H., Bournazou E., Schori E., Zhang H., Brewer M.R., Pao W., Morris L., Ladanyi M., Arcila M., Manova-Todorova K., de Stanchina E., Norton L., Levine R.L., Altan-Bonnet G., Solit D., Zinda M., Huszar D., Lyden D, & Bromberg J.F. (2016). JAK2 inhibition sensitizes resistant EGFR-mutant lung adenocarcinoma to tyrosine kinase inhibitors. Science signaling, 9(421), ra33.

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Antibody Characterization for Cell Signaling

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Primary antibodies used for western blot experiments: phospho-p53 Ser15, cleaved Caspase-3 and cleaved PARP (Cell Signaling, Leiden, The Netherlands), MAPK15 (custom preparation), p53 (Santa Cruz Biotechnology), ACTB (Sigma Aldrich), CDKN1A/p21 (Epitomics, Burlingame, CA), LC3B (Novus Biologicals, Cambridge, UK), SQSTM1/p62 (BD Biosciences, Milan, Italy), GADD45a (Cell Signaling), phospho-ATR (Cell Signaling), phospho-CHK2 (Cell Signaling), PCNA (Cell Signaling). Primary antibodies used for immunofluorescence experiments: γH2A.X (Cell Signaling), 53BP1 (Novus Biologicals) LC3B (MBL, Woburn, MA). Primary antibody used for immunohistochemistry experiments: MAPK15 (custom preparation). Secondary antibodies used for western blot experiments: HRP-conjugated anti-Mouse IgG and HRP-conjugated anti-Rabbit IgG (Santa Cruz Biotechnology). Secondary antibodies used for immunofluorescence experiments: AlexaFluor488-conjugated anti-Rabbit IgG and AlexaFluor555-conjugated anti-Mouse IgG (Life Technologies).

Rossi M., Colecchia D., Ilardi G., Acunzo M., Nigita G., Sasdelli F., Celetti A., Strambi A., Staibano S., Croce C.M, & Chiariello M. (2016). MAPK15 upregulation promotes cell proliferation and prevents DNA damage in male germ cell tumors. Oncotarget, 7(15), 20981-20998.

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VEGF Protein Quantification by Western Blot

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VEGF protein levels were detected by Western blot, using 30 μg of protein in 8.5% SDS-PAGE electrophoresis gel and electroblotted. The membranes were incubated with mouse anti-VEGF antibody (1:1000, ThermoScientific) overnight. Next, the membranes were incubated with a secondary antibody and HRP-conjugated anti-mouse IgG (1:5000, Santa Cruz Biotechnology). The proteins were detected with an enhanced chemiluminescence kit (Millipore) and by radiography. All Western blot analyses were performed within the linear range of protein loads and antibody use. The bands were scanned for densitometric analysis using the UVP EC3 Imaging System and the UVP VisionWorks LS Image acquisition and Analysis Software.

Lima-Posada I., Portas-Cortés C., Pérez-Villalva R., Fontana F., Rodríguez-Romo R., Prieto R., Sánchez-Navarro A., Rodríguez-González G.L., Gamba G., Zambrano E, & Bobadilla N.A. (2017). Gender Differences in the Acute Kidney Injury to Chronic Kidney Disease Transition. Scientific Reports, 7, 12270.

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Protein Expression Analysis Protocol

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The cells were lysed in RIPA with 1 mM PMSF and protease inhibitors (Sigma) for 15 min on ice. The cytosol and nuclear fraction were isolated according to the manufactory's instruction (Thermo). Similar amounts of protein from each extract were subjected to SDS-PAGE analysis and transferred to polyvinyl difluoride (PVDF) membranes (Millipore). After blocking for 1 h with blocking buffer (5% nonfat milk and 0.1% Tween-20 in PBS), the membranes were incubated with the following primary antibodies at 4°C overnight: anti-phospho-p38 MAPK, anti-p38 MAPK, anti-phospho-JNK, anti-JNK (Santa Cruz), anti-phospho-ERK, anti-ERK, anti-phospho-Akt, anti-Akt, anti-phospho-IκBα, anti-IκBα, anti-p50/p105, anti-p65, anti-Histone H3 (Cell Signaling Technology), anti-gC1qR (Hycult biotech) and anti-β-actin (Tianjin Sungene Biotech) antibodies. HRP-conjugated anti-mouse IgG or anti-rabbit IgG (Santa Cruz) were used as secondary antibodies. And ECL (Bio-Rad) was used for chemiluminescent detection according to the manufacturer's instructions.

Du Q., Huang Y., Wang T., Zhang X., Chen Y., Cui B., Li D., Zhao X., Zhang W., Chang L, & Tong D. (2016). Porcine circovirus type 2 activates PI3K/Akt and p38 MAPK pathways to promote interleukin-10 production in macrophages via Cap interaction of gC1qR. Oncotarget, 7(14), 17492-17507.

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Western Blot Analysis of Soluble and Insoluble Brain Fractions

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WB with soluble and insoluble fractions of four brain homogenates were performed as described in10 (link)44 (link)55 (link)56 (link). Briefly, soluble and insoluble fractions applied to electrophoresis on NuPAGE 4–12% Bis-Tris gel in MES buffer under reducing conditions (Invitrogen, CA) and electrotransferred onto nitrocellulose membrane (GE Healthcare, NJ). For antigen retrieval, membranes with soluble fractions were boiled in PBS at 100 °C as described in Rosen et al.58 (link) and then were blocked with 5% fat-free dry milk. Membranes with insoluble fractions were blocked without antigen retrieval. Aβ and tau were visualized by incubating with sera from mice immunized with AV-1959R, AV-1980R, AV-1953R or mixture of AV-1959R and AV-1980R followed by HRP-conjugated anti-mouse IgG (Santa Cruz Biotechnology, CA). Sera were used after normalization of antibody concentration measured by ELISA (1 μg/ml for anti-Aβ and 0.4 μg/ml for anti-tau).

Davtyan H., Zagorski K., Rajapaksha H., Hovakimyan A., Davtyan A., Petrushina I., Kazarian K., Cribbs D.H., Petrovsky N., Agadjanyan M.G, & Ghochikyan A. (2016). Alzheimer’s disease AdvaxCpG- adjuvanted MultiTEP-based dual and single vaccines induce high-titer antibodies against various forms of tau and Aβ pathological molecules. Scientific Reports, 6, 28912.

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Quantitative Analysis of Tau Protein in AD

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Preparation of brain homogenates, Western blot (WB) and Dot blot (DB) analysis were performed as previously described^{30 (link),62 (link)}. Briefly, 0.2 g of brain tissue from four different AD cases were homogenized in 0.4 ml TBS buffer with Halt™ Protease and Phosphatase Inhibitor Cocktail (100X, Thermo Scientific, CA), then centrifuged at 6400xg for 15 minutes at +4 °C. Supernatants (soluble fractions) were collected and stored at −80 °C for further analysis. For WB soluble fractions applied to electrophoresis on NuPAGE 4–12% Bis-Tris gel in MES buffer under reducing conditions (Invitrogen, CA) and electrotransferred onto nitrocellulose membrane (GE Healthcare, NJ). Tau were visualized by incubating with sera (dilution at 1:1000) from mice immunized with AV-1980R/A and injected with Advax^CpG only followed by HRP-conjugated anti-mouse IgG (Santa Cruz Biotechnology, CA). For DB assay the same extracts were applied to membrane (1 μg). Proteins were detected using sera from mice immunized with AV-1980R/A and control mice injected with Advax^CpG only, TNT-1 (Millipore, MA), HT7 (Life Technology, CA) antibodies. All primary antibodies were used at concentration of 1 μg/ml, serum was used at dilution 1:2500. Bovine anti-mouse HRP-conjugated secondary antibody was used (Santa Cruz Biotechnology, CA).

Hovakimyan A., Antonyan T., Shabestari S.K., Svystun O., Chailyan G., Coburn M.A., Carlen-Jones W., Petrushina I., Chadarevian J.P., Zagorski K., Petrovsky N., Cribbs D.H., Agadjanyan M.G., Ghochikyan A, & Davtyan H. (2019). A MultiTEP platform-based epitope vaccine targeting the phosphatase activating domain (PAD) of tau: therapeutic efficacy in PS19 mice. Scientific Reports, 9, 15455.

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RAW264.7 Cell Protein Expression Analysis

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RAW264.7 cells were seeded in 6-well plates. The cells were treated with 50, 100, or 200 μg/mL of TVE for 1 h and sensitized with LPS (1 µg/mL) for another 24 h or 15 min of incubation. The cells were subsequently harvested and washed twice with ice PBS (phosphate buffered saline) and pelleted by centrifugation for 20 min at 12,000 rpm. The protein concentration was measured using a BCA^TM protein assay kit (Thermo Scientific, Waltham, MA, USA) according to manufacturer’s instructions. β-actin, inducible NO synthase [iNOS], cyclooxygenase-2 [COX-2], Ik-B, phospho (p)-p65, ERK, p-ERK, JNK, p-JNK, p-38, and p-p38 were used as the primary antibodies (1:1000 dilution, Cell Signaling Technology, Beverly, MA, USA). The HRP-conjugated anti-mouse IgG and anti-rabbit IgG were used secondary antibodies (1:5000 dilution, Santa Cruz Biotechnology, CA, USA). A densitometric analysis was performed using ImageJ to quantify protein expression [25 (link)].

Kim E.A., Kang N., Kim J., Yang H.W., Ahn G, & Heo S.J. (2022). Anti-Inflammatory Effect of Turbo cornutus Viscera Ethanolic Extract against Lipopolysaccharide-Stimulated Inflammatory Response via the Regulation of the JNK/NF-kB Signaling Pathway in Murine Macrophage RAW 264.7 Cells and a Zebrafish Model: A Preliminary Study. Foods, 11(3), 364.

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Protein Expression Analysis of FOSL1 and FOSL2

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Cell culture pellets
were lysed using radioimmunoprecipitation assay buffer (Pierce, cat
no. 89901) that was supplemented with protease and phosphatase inhibitors
(Roche) and sonicated using a Bioruptor UCD-200 (Diagenode). Sonicated
lysates were centrifuged at 14,000 rpm for 30 min at 4 °C, and
supernatants were collected. Samples were estimated for protein concentration
(DC protein assay; Bio-Rad) and boiled in 6× Laemmli buffer (330
mM Tris-HCl, pH 6.8; 330 mM sodium dodecyl sulfate; 6% β-ME;
170 μM bromophenol blue; and 30% glycerol). Samples were then
loaded on gradient Mini-PROTEAN TGX precast protein gels (Bio-Rad)
and transferred to polyvinylidene fluoride membranes (Trans-Blot Turbo
transfer packs, Bio-Rad).
For protein expression analysis of
FOSL1 and FOSL2, the following Ab were used: anti-FOSL1 (Cell Signaling
Tech., cat no. 5281), anti-FOSL2 (Cell Signaling Tech., cat no. 19967),
and anti-β-actin (Sigma-Aldrich, cat no. A5441). HRP-conjugated
anti-mouse IgG (Santa Cruz Biotechnology, cat no. sc-2005) and anti-rabbit
IgG (BD PharMingen, cat no. 554021) were used as secondary Ab.

Shetty A., Bhosale S.D., Tripathi S.K., Buchacher T., Biradar R., Rasool O., Moulder R., Galande S, & Lahesmaa R. (2021). Interactome Networks of FOSL1 and FOSL2 in Human Th17 Cells. ACS Omega, 6(38), 24834-24847.

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