Hrp conjugated anti mouse igg

Manufactured by Merck Group

Sourced in United States, Germany

HRP-conjugated anti-mouse IgG is a laboratory reagent used for the detection of mouse immunoglobulin G (IgG) in various immunoassay applications. It consists of a horseradish peroxidase (HRP) enzyme conjugated to an antibody that specifically binds to mouse IgG. This product can be used to facilitate the visualization and quantification of mouse IgG in samples.

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Lab products found in correlation

Hrp conjugated anti rabbit igg, by Merck Group (21 mentions) Anti β actin, by Merck Group (5 mentions) Pvdf membrane, by Merck Group (3 mentions) 3 3 diaminobenzidine (dab), by Merck Group (3 mentions) Hybond ecl, by GE Healthcare (3 mentions) Cy3 conjugated anti rabbit igg, by Thermo Fisher Scientific (3 mentions) Alexaflour 488 conjugated anti mouse igg, by Thermo Fisher Scientific (3 mentions) Nitrocellulose membrane, by Merck Group (3 mentions) Anti β catenin, by BD (3 mentions) Nunc immuno plate, by Thermo Fisher Scientific (3 mentions) Ripa buffer, by Solarbio (2 mentions) β actin, by Cell Signaling Technology (2 mentions) Monoclonal anti β actin, by Merck Group (2 mentions) Clarity and clarity max ecl, by Bio-Rad (2 mentions) Mouse anti v5, by Thermo Fisher Scientific (2 mentions) Chemidoc mp system, by Bio-Rad (2 mentions) Enhanced chemiluminescent kit, by CWBIO (2 mentions) 3 3 diaminobenzidine tetrahydrochloride dab, by Merck Group (2 mentions) Ecl reagent kit chemiluminescence substrate, by GE Healthcare (2 mentions) 3 3 5 5 tetramethylbenzidine (tmb), by Merck Group (2 mentions)

117 protocols using hrp conjugated anti mouse igg

Screening and Characterizing Anti-ZIKV mAbs

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To screen hybridomas, micro-ELISA plates (Nunc, USA) were coated overnight at 4 °C with 200 ng/well of ZIKV E80 protein, and blocked with 5% milk in PBS-Tween20 (PBST). 50 μL of undiluted hybridoma culture supernatants was added to the plates and incubated at 37 °C for 2 h. Plates were then washed with PBST and incubated with horseradish peroxidase (HRP)-conjugated anti-mouse IgG (Sigma, USA). After washes and color development, absorbance at 450 nm was measured.
Immunoglobulin isotypes of the mAbs were measured using SBA Clonotyping^TM System/HRP ELISA kit (Southern Biotech, USA) according to manufacturer’s instructions.
To measure binding properties of these mAbs, microplates (Nunc) were coated at 4 °C overnight with 200 ng/well of ZIKV E80, or DENV2 E80, and then blocked with 5% milk in PBST. Next, 50 μL/well of serially diluted anti-ZIKV mAbs, anti-EV71 mAb D5 (isotype control)^{35 (link)} or anti-DENV mAb D1-11 (Santa Cruz Biotechnology, USA) were added and incubated at 37 °C for 2 h. After washing with PBST, plates were incubated with HRP-conjugated anti-mouse IgG (Sigma). After color development, absorbance at 450 nm was measured.

Qu P., Zhang C., Li M., Ma W., Xiong P., Liu Q., Zou G., Lavillette D., Yin F., Jin X, & Huang Z. (2020). A new class of broadly neutralizing antibodies that target the glycan loop of Zika virus envelope protein. Cell Discovery, 6, 5.

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Purification and Analysis of Diubiquitin Binding

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His pull-down assays were performed as described in [6 , 45 (link)]. Briefly, 500 pmol of purified Histagged hRpn10^196–306 was added to 20 μL of pre-washed Ni-NTA agarose resin for one hour and washed once with Buffer 1 (50 mM HEPES, 50 mM NaCl, 5% (v/v) glycerol, and 15 mM 2 mercaptoethanol at pH 6.7). The resin was then incubated with 500 pmol of M1, K6, K11, K27, K29, K33, K48, and K63 diubiquitin for one hour. Intrinsically disordered protein SocB with a C-terminal His-tag [80 (link)] was used as a negative control with K48 diubiquitin. Unbound protein was removed by extensive washing in the above buffer. The resin-bound proteins were fractionated by electrophoresis and transferred to a PVDF membrane. The membrane was treated with denaturing Buffer 2 (20 mM Tris-HCl, 6 M guanidine hydrochloride, 1 mM PMSF, and 5 μM 2 -mercaptoethanol at pH 7.4) for 30 mins at 4°C, extensively washed with Trisbuffered saline +0.1% Tween, and analyzed by immunoblotting. Diubiquitin was detected with mouse anti-Ub antibody (Millipore Sigma MAB1510, 1:1000) followed by HRP-conjugated anti-mouse IgG (Millipore Sigma, 1:5000). His-tagged hRpn10^196–306 or SocB-His was detected with mouse anti-His antibody (Thermo Fisher Scientific MA1–21315, 1:1000) followed by HRP-conjugated anti-mouse IgG (Millipore Sigma, 1:5000).

Chen X., Ebelle D.L., Wright B.J., Sridharan V., Hooper E, & Walters K.J. (2019). Structure of hRpn10 bound to UBQLN2 UBL illustrates basis for complementarity between shuttle factors and substrates at the proteasome. Journal of molecular biology, 431(5), 939-955.

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Generating Dog BC Organoids for Drug Screening

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To generate dog BC organoids, cells from urine samples were mixed with Matrigel (BD Bioscience) and cultured with stem cell‐stimulated medium, as described previously.17 Anticancer drugs used were as follows: piroxicam; gemcitabine; vinblastine (Cayman); and cisplatin (WAKO). Antibody sources used were as follows: E‐cadherin (R&D System); CK7; Ki67 (Novus); CK20; UPK3; MMP28; TFPI2; AGPAT4 (Bioss); vimentin (Sigma‐Aldrich); α‐smooth muscle actin (SMA) (DAKO); CK5; CNN3 (GeneTex, Inc.); and CTSE (Bioworld Technology, Inc.). Fluorescent secondary antibodies used were as follows: Alexa Fluor™ 488 donkey anti‐goat IgG; Alexa Fluor 488™ goat anti‐rabbit IgG; Alexa Fluor 488™ goat anti‐mouse IgG; (Thermo Fisher Scientific Inc.); Biotinylated goat anti‐mouse IgG (Vector Laboratories, Inc.). Horseradish peroxidase (HRP)‐conjugated anti‐rabbit IgG (Cayman); and HRP‐conjugated anti‐mouse IgG (Millipore).

Elbadawy M., Usui T., Mori T., Tsunedomi R., Hazama S., Nabeta R., Uchide T., Fukushima R., Yoshida T., Shibutani M., Tanaka T., Masuda S., Okada R., Ichikawa R., Omatsu T., Mizutani T., Katayama Y., Noguchi S., Iwai S., Nakagawa T., Shinohara Y., Kaneda M., Yamawaki H, & Sasaki K. (2019). Establishment of a novel experimental model for muscle‐invasive bladder cancer using a dog bladder cancer organoid culture. Cancer Science, 110(9), 2806-2821.

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Western Blot Analysis of Cell Cycle Proteins

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Proteins were extracted using RIPA buffer (Solarbio, Beijing, China, R0020) according to the manufacturer’s protocol. The concentration of protein was measured by BCA kit (Takara, Japan, AI90451A). Thirty micrograms of total proteins from each sample were separated by polyacrylamide gel electrophoresis and transferred onto poly-vinylidene fluoride (PVDF) membranes (Millipore, Billerica, MA, USA). The blot-transferred membranes were blocked with 5% fat-free dry milk for 2 h and incubated with primary antibodies: mouse anti-PCNA (1:500; Santa Cruz Biotechnology, PC10), mouse anti-CDK1 (1:500; Santa Cruz Biotechnology, AN21.2), mouse anti-CDK2 (1:500; Santa Cruz Biotechnology, D-12), and mouse anti beta-ACTIN (1:2000; CWBIO, Beijing, China, CW0096) on a shaker at 4 °C overnight, followed by incubation with horseradish peroxidase (HRP)–conjugated anti-mouse IgG (1:2000; Millipore, AP192P). Protein bands were visualized and captured by a Bio-Rad Chemidoc XRS using a Western Bright ECL Kit (Bio-Rad, Berkeley, CA, USA). The protein bands were hand-drawn for 3 times and calculated the average intensity by Image-Pro Plus. The ratio of the target protein to the corresponding intensity value of β-actin was obtained for analysis. Experiments were repeated three times.

Li T., Chen Q., Zheng Y., Zhang P., Chen X., Lu J., Lv Y., Sun S, & Zeng W. (2019). PAMAM-cRGD mediating efficient siRNA delivery to spermatogonial stem cells. Stem Cell Research & Therapy, 10, 399.

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Western Blot Analysis of Protein Samples

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Total extracts of various cells were obtained after lysis for 30 min at 4 °C in RIPA buffer (150 mM NaCl, 1% NP-40, 0.5% Doxycholic acid, 0.1% SDS, 50 mM Tris-HCl (pH 7.4)). Protein samples were fractionated by sodium dodecyl-sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) on a 12% polyacrylamide gel under denaturing conditions and transferred to a nitrocellulose membrane. The membrane was blocked with 5% skim milk in PBST (PBS containing 0.1% Tween 20) at room temperature (RT) for 2 hrs. After washing with PBST, the membrane was incubated with various primary antibodies at RT for 1 hr, followed by horseradish peroxidase (HRP)-conjugated anti-mouse IgG (Millipore). The stained bands were visualized by using ECL (Animal Genetics, Gyeonggi-do, Korea) detection reagent.

Kim J.Y., Kim S.Y., Choi H.S., An S, & Ryu C.J. (2019). Epitope mapping of anti-PGRMC1 antibodies reveals the non-conventional membrane topology of PGRMC1 on the cell surface. Scientific Reports, 9, 653.

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Immunoblot Analysis of VeA and LaeA

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Western blot analyses of VeA and LaeA were performed as described in Jeong and Yu (44 (link)). Briefly, 2-day-old conidia (2 × 10⁸), vegetatively grown cells (6, 12, 24, 48 h), asexually developing cells (12, 24, 48 h), and sexually developing cells (12, 24, 48 h) were collected and resuspended in the lysis buffer containing a 1× protease inhibitor cocktail. Then, samples were homogenized by a mini-beadbeater with 0.5 mm zirconia/silica beads. Protein concentrations were measured by the Bio-Rad Protein Assay (Bio-Rad). Approximately 15 μg of total proteins per lane were separated on 4 to 12% gradient SDS-PAGE (Bio-Rad) gel and transferred onto immobilon-P PVDF membrane (Millipore). The membrane was blocked with a blocking buffer, incubated with the monoclonal anti-FLAG antibody produced in mice (clone M2, Sigma-Aldrich), and then incubated with HRP-conjugated anti-mouse IgG (Millipore). The membrane was developed using Amersham enhanced chemiluminescence detection reagents (GE Healthcare).

Moon H., Lee M.K., Bok I., Bok J.W., Keller N.P, & Yu J.H. (2023). Unraveling the Gene Regulatory Networks of the Global Regulators VeA and LaeA in Aspergillus nidulans. Microbiology Spectrum, 11(2), e00166-23.

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Colorectal Tumor-Derived Organoid Culture

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Human colorectal tumor tissue-derived ALI organoids were cultured as described previously [10 (link)]. The medium components were as follows: Advanced Dulbecco’s Modified Eagle’s Medium (DMEM) with 50% Wnt, Noggin and R-Spondin conditioned medium; GlutaMax; B-27 supplement; 100 μg/mL Primocin (Invitrogen, Carlsbad, CA, USA); 1 mM N-Acetyl-l-cysteine; 10 mM Nicotinamide (Sigma-Aldrich, St. Louis, MO, USA); 50 ng/mL mouse epidermal growth factor (EGF) (PeproTech, Inc., Rocky Hill, NJ, USA); 500 nM A83-01 (Adooq Bioscience, Irvine, CA, USA); and 3 μM SB202190; 10 μM Y-27632 (Cayman, Ann Arbor, MI, USA). Stem cell-related signal inhibitors were as follows: YO-01027 (Toronto Research Chemicals, Toronto, Canada); DAPT (Adooq Bioscience); WAV939; Wnt-C59; AY9944; and GANT61 (Cayman). Anti-cancer drugs were as follows: 5-FU (WAKO, Tokyo, Japan); Irinotecan (LC Laboratories, Woburn, MA, USA); and Oxaliplatin (Adooq Bioscience). Antibody sources were as follows: GLI-1 (Gene Tex, Irvine, CA, USA); CD44 (Bethyl Laboratories, Montgomery, TX, USA); c-Myc; and Nanog (Cell Signaling, Beverly, MA, USA). Secondary antibodies were as follows: Horseradish peroxidase (HRP) conjugated anti-rabbit IgG; HRP conjugated anti-goat IgG (Cayman); and HRP conjugated anti-mouse IgG (Millipore, Temecula, CA, USA).

Usui T., Sakurai M., Umata K., Elbadawy M., Ohama T., Yamawaki H., Hazama S., Takenouchi H., Nakajima M., Tsunedomi R., Suzuki N., Nagano H., Sato K., Kaneda M, & Sasaki K. (2018). Hedgehog Signals Mediate Anti-Cancer Drug Resistance in Three-Dimensional Primary Colorectal Cancer Organoid Culture. International Journal of Molecular Sciences, 19(4), 1098.

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Western Blot Analysis of Protein Markers

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Total protein (50 μg) from samples was separated by 12% SDS-PAGE and transferred to 0.22 micrometer Immobilon-P membranes(Millipore). The membranes were incubated overnight at 4°C with anti-HPV16 E7(Santa Cruz) diluted at 1:500, anti-human p53(Millipore) diluted at 1:1000, anti-human PLK2(Proteintech) diluted at 1:1000, anti-human DGCR8(Proteintech) diluted at 1:1000 or β-actin(Cell Signaling) diluted at 1:1000. After washed with PBST for three times, the membranes were incubated with horseradish peroxidase (HRP)-conjugated anti-mouse IgG (Millipore) diluted at 1:5000 for two hours. HRP activity was detected with SuperSignal West Pico enhanced chemiluminescent (ECL) substrate (Thermo Scientific).

Liu F., Zhang S., Zhao Z., Mao X., Huang J., Wu Z., Zheng L, & Wang Q. (2016). MicroRNA-27b up-regulated by human papillomavirus 16 E7 promotes proliferation and suppresses apoptosis by targeting polo-like kinase2 in cervical cancer. Oncotarget, 7(15), 19666-19679.

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Phosphorylation of Pyruvate Dehydrogenase

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Cells washed with ice-cold PBS, and scraped into cold RIPA buffer containing cOmplete Mini protease inhibitor (Roche, 11836170001) and PhosStop Phosphatase Inhibitor Cocktail Tablets (Roche, 04906845001). Protein concentration was calculated using the BCA Protein Assay (Pierce, 23225) with BSA as a standard. Lysates were resolved by SDS-PAGE and proteins were transferred onto nitrocellulose membranes using the iBlot2 Dry Blotting System (Thermo Fisher, IB21001, IB23001). Protein was detected with the primary antibodies anti-Pyruvate Dehydrogenase E1-alpha subunit (phospho S293) (Abcam, ab92696), anti-PDK1 (Cell Signaling Technologies, 3062S), anti-FLAG (Sigma, F1804) and anti-Vinculin (Sigma, V9131). The secondary antibodies used were IR680LT dye conjugated anti-rabbit IgG (Licor Biosciences, 925-68021), IRDye 800CW conjugated anti-mouse IgG (Licor Biosciences, 925-32210), HRP-conjugated anti-rabbit IgG (Millipore, 12-348), and HRP-conjugated anti-mouse IgG (Millipore, 12-349).

Luengo A., Li Z., Gui D.Y., Sullivan L.B., Zagorulya M., Do B.T., Ferreira R., Naamati A., Ali A., Lewis C.A., Thomas C.J., Spranger S., Matheson N.J, & Vander Heiden M.G. (2020). Increased demand for NAD+ relative to ATP drives aerobic glycolysis. Molecular cell, 81(4), 691-707.e6.

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Immunohistological Analysis of MHC Expression

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Before the immunohistological analysis, tissue sections were deparaffinized with xylene and rehydrated through ethanol exchange. Antigen retrieval was performed on the sections with EDTA buffer (pH = 8) in the stream chamber for 30 min. Endogenous peroxidase activity was blocked with 0.3% hydrogen peroxide for 20 min at room temperature. Before the blocking step, sections were washed with 0.025% Triton X-100 in TBS. Sections were then blocked with 10% Normal Goat Serum (NGS) in 1% BSA in TBS for 1 h at room temperature followed by incubation with mouse anti-MHC (1:50, MAB4470, R&D Systems) antibody overnight at 4 °C. Sections were then washed with 0.025% Triton X-100 in TBS and incubated with a secondary antibody (HRP conjugated antimouse IgG, 1:500, 12-349, Millipore) for 1 h at room temperature. Both primary and secondary antibodies are prepared with 1% BSA in TBS. Sections were washed with 0.025% Triton X-100 in TBS and incubated with DAB solution for 20 min. Sections were then counterstained in hematoxylin and dehydrated.

Eren Cimenci C., Uzunalli G., Uysal O., Yergoz F., Karaca Umay E., Guler M.O, & Tekinay A.B. (2017). Laminin mimetic peptide nanofibers regenerate acute muscle defect. Acta biomaterialia, 60.

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