Mouse igg blocking reagent

Manufactured by Vector Laboratories

Sourced in United States

The Mouse IgG blocking reagent is a solution designed to block non-specific binding of mouse immunoglobulin G (IgG) in immunohistochemistry and other immunoassay applications. It helps minimize background signal by preventing unwanted interactions between the sample's endogenous mouse IgG and the detection system.

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Lab products found in correlation

Alexa fluor 488 phalloidin, by Thermo Fisher Scientific (2 mentions) Tcs sp2 spectral confocal microscope, by Leica (2 mentions) Anti pericentrin antibody, by Abcam (2 mentions) Anti rabbit cy3, by Jackson ImmunoResearch (2 mentions) Hoechst 33342, by Thermo Fisher Scientific (2 mentions) Ab9610, by Merck Group (1 mentions) Alexa fluor 488 conjugated donkey anti rabbit, by Thermo Fisher Scientific (1 mentions) Ab7349, by Abcam (1 mentions) Af1062, by R&D Systems (1 mentions) Bz 9000, by Keyence (1 mentions) Gelatin, by Merck Group (1 mentions) Hydromount, by National Diagnostics (1 mentions) Bsa c, by Aurion (1 mentions) Bovine serum albumin (bsa), by Aurion (1 mentions) Bz x analyzer, by Keyence (1 mentions) Bz x710 microscope, by Keyence (1 mentions) Tyramide alexa fluor 488, by Thermo Fisher Scientific (1 mentions) Anti human cd45, by Agilent Technologies (1 mentions) Discovery xt system, by Roche (1 mentions) Mom kit bmk 2202, by Vector Laboratories (1 mentions)

11 protocols using mouse igg blocking reagent

Oligodendrocyte Immunohistochemistry in CNS

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The corpus callosum coronal sections and the spinal cord transverse sections were immunostained to evaluate oligodendrocytes. These sections were permeabilized with 0.3% Triton X-100, and nonspecific binding sites were blocked with serum matching the species of the secondary antibody. Where anti-mouse IgG monoclonal antibodies were used, sections were additionally incubated with mouse IgG blocking reagent (Vector Laboratories, Burlingame, CA). The sections were then stained with the following primary antibodies: rabbit anti-Olig-2 (1 : 500, AB9610, Sigma-Aldrich, St. Louis, MO, RRID:AB_570666), goat anti-platelet-derived growth factor receptor alpha (PDGFRα, 1 : 200, AF1062, R&D Systems, Minneapolis, MN, RRID:AB_2236897), mouse anti-CC1 (1 : 100, OP80, Sigma-Aldrich, RRID:AB_2057371), and rat anti-myelin basic protein (MBP, 1 : 250, ab7349, Abcam, Cambridge, UK, RRID:AB_305869). The following secondary antibodies (1 : 500, Thermo Fisher Scientific) were used: Alexa Fluor 350-conjugated donkey anti-goat, Alexa Fluor 488-conjugated donkey anti-rabbit, Alexa Fluor 594-conjugated donkey anti-mouse, Alexa Fluor 594-conjugated goat anti-rat, and Alexa Fluor 594-conjugated goat anti-rabbit. Immunostained sections were observed under a fluorescence microscope (BZ9000; Keyence, Osaka, Japan).

Tsuchikawa Y., Kamei N., Sanada Y., Nakamae T., Harada T., Imaizumi K., Akimoto T., Miyaki S, & Adachi N. (2023). Deficiency of MicroRNA-23-27-24 Clusters Exhibits the Impairment of Myelination in the Central Nervous System. Neural Plasticity, 2023, 8938674.

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Immunohistochemical Analysis of eNOS and 8-OHdG

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Paraffin‐embedded sections were deparaffinized in xylene and rehydrated in 100%, 90%, 80%, 70%, and 50% ethanol. Microwave treatment was done to activate the enzymes. To reduce the background staining, nonspecific binding was blocked by incubating with blocking solution [PBS (pH 7.2) containing 2% bovine serum albumin, 2% fetal bovine serum, and 0.2% fish gelatin, added mouse IgG blocking reagent (Vector Laboratories)]. The sections were then incubated with the primary antibody [purified mouse anti‐endothelial nitric oxide synthase (eNOS)/NOS Type 3, BD Transduction Laboratories, San Jose, NJ, USA, and anti‐8‐hydroxy‐2′‐deoxyguanosine (8‐OHdG) monoclonal antibody, Japan Institute for the Control of Aging, Shizuoka, Japan] diluted 1 : 100 in blocking solution at 4 °C. Peroxidase activity was developed in 3,3‐diaminobenzidine. Finally, Mayer's hematoxylin was added as a counterstain. The secondary antibody was anti‐mouse Envision Plus polymer reagent (DAKO, Santa Clara, CA, USA).

Ichikawa S., Gohda T., Murakoshi M., Li Z., Adachi E., Koshida T, & Suzuki Y. (2020). Aspartic acid supplementation ameliorates symptoms of diabetic kidney disease in mice. FEBS Open Bio, 10(6), 1122-1134.

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Quantifying Centrosome Dynamics in Hepatocytes

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Liver cryosections (5μm) were fixed with 4% paraformaldehyde and blocked for 1 hour with mouse IgG blocking reagent (vector labs). After blocking with Donkey serum (5% in PBS-0.3%Tween), sections were incubated with 1/500 anti-pericentrin antibody (Abcam) overnight. Secondary 1/500 Anti-rabbit Cy3 (Jackson immunoresearch, stratech) and 1/500 Alexafluor 488 –phalloidin (invitrogen) was then applied on the sections for 1-2hours. Sections were mounted with vectashield plus DAPI medium. Sections were viewed and analysed using a Leica TCS SP2 spectral confocal microscope and Leica confocal software (Leica, Heidelberg, Germany). Centrosomes numbers were counted in at least 150 nuclei of hepatocytes from each mouse for each genotype (N=3-5) at D4 and D6.

Méniel V., Megges M., Young M.A., Cole A., Sansom O.J, & Clarke A.R. (2014). Apc and p53 interaction in DNA damage and genomic instability in hepatocytes. Oncogene, 34(31), 4118-4129.

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Quantifying Centrosome Dynamics in Hepatocytes

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Méniel V., Megges M., Young M.A., Cole A., Sansom O.J, & Clarke A.R. (2014). Apc and p53 interaction in DNA damage and genomic instability in hepatocytes. Oncogene, 34(31), 4118-4129.

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Coronal Brain Section Immunostaining

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30μm coronal hemi-brain sections were washed 2×10min in phosphate-buffered saline (PBS) then exposed to UV light overnight to reduce autofluorescence. The next day, slices were washed 3×10min in PBS, then washed 2×15min in PBS + 0.1% Tween-20 (PBS-T). Slices were blocked in PBS + 10% Normal Donkey Serum + 0.2% Gelatin (Sigma) + 0.5% Triton-X for 1hr at room temperature (RT), then washed for 10min in PBS. Because some antibodies were raised in mouse, slices were additionally blocked in 1 drop Mouse IgG Blocking Reagent (Vector Labs, MKB-2213-1) per 5mL PBS for 1 hour at RT. Primary antibodies were diluted to optimized concentrations (anti-NeuN 1:1000, anti-MHC-I (OX18 that recognizes multiple MHC-I proteins) 1:50, anti-PSD-95 1:200) in 1:12.5 mouse-on-mouse Protein Concentrate (Vector Labs, MKB-2213-1) in PBS, and slices were incubated overnight at 4°C. The next day, slices were washed 3x (15 min, 10 min, 5 min) in PBS-T. Secondary antibodies (Lifetech, Jackson ImmunoResearch; 1:1000) were diluted in the same dilution buffer as primary antibodies and incubated 1hr at RT. Slices were then washed 3x (15 min, 10 min, 5 min) with PBS, mounted with Vectashield + DAPI, and coverslipped.

Zalocusky K.A., Najm R., Taubes A.L., Hao Y., Yoon S.Y., Koutsodendris N., Nelson M.R., Rao A., Bennett D.A., Bant J., Amornkul D.E., Xu Q., An A., Cisne-Thomson O, & Huang Y. (2021). Neuronal ApoE Upregulates MHC-I Expression to Drive Selective Neurodegeneration in Alzheimer’s Disease. Nature neuroscience, 24(6), 786-798.

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Immunohistochemical Analysis of Intestinal Tissues

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Sections were cut at 10 μm and randomly oriented on the slide. Five cryosections at 50 μm intervals from each mouse intestine were used for staining. Cryosections were dried for 20 minutes then fixed in 4% paraformaldehyde (PFA) for 20 minutes and rinsed in phosphate-buffered saline (PBS) for 2 minutes at room temperature. All sections were subjected to immunohistochemical analysis using primary and secondary antibodies listed in Table 1.
For PGP9.5 primary antibody staining, sections were subjected to antigen retrieval by boiling slides in 20mM citrate buffer (Thermo Scientific) for 15 minutes, and blocked in PBS/M.O.M (mouse IgG blocking reagent, Vector) for 1 hour at room temperature. After incubation with primary antibodies at room temperature for 1 hour, slides were rinsed in PBS/0.03% Triton for 2 minutes twice followed by incubation in secondary antibody for 1 hour. Slides were then rinsed twice with PBS/0.03% Triton for 2 minutes and mounted with hydromount (National Diagnostics) and DAPI (1:1500, Hoechst 33342, ThermoFisher Scientific) for nuclear staining.

Sintusek P., Catapano F., Angkathunkayul N., Marrosu E., Parson S.H., Morgan J.E., Muntoni F, & Zhou H. (2016). Histopathological Defects in Intestine in Severe Spinal Muscular Atrophy Mice Are Improved by Systemic Antisense Oligonucleotide Treatment. PLoS ONE, 11(5), e0155032.

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Immunohistochemical Analysis of Transplanted Testes

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Testes of transplanted and control mice were fixed, dehydrated, embedded in paraffin and serial sectioned (5 μm), and every fourth section was used for immunohistochemical analysis. In short, after removal of the paraffin, we performed heat-induced antigen retrieval (10 minutes) using 0.01 M tri-sodium citrate dehydrate buffer (pH = 6). Sections were blocked for 1 h with mouse IgG blocking reagent (Vector Laboratories, USA), after which we blocked with 5% BSA + 0.5% BSAc (Aurion, the Netherlands) for 1 h. Sections were incubated with the anti-human nucleoli primary antibody (Supplementary Table SI) overnight in a humidified box at 4°C. Endogenous peroxidase was blocked with 3% H₂O₂ (Millipore, USA) in methanol. As a secondary antibody, poly-HRP Goat-anti-Mouse/Rabbit (Immunologic, the Netherlands) was used with 30 minutes incubation at room temperature. The DAB bright kit (Immunologic, the Netherlands) was used to visualize the reaction product, followed by counterstaining with hematoxylin. The researcher who analyzed the sections was blinded for the origin of the sections (transplanted or not-transplanted tissue).

Eliveld J., van den Berg E.A., Chikhovskaya J.V., van Daalen S.K., de Winter-Korver C.M., van der Veen F., Repping S., Teerds K, & van Pelt A.M. (2019). Primary human testicular PDGFRα+ cells are multipotent and can be differentiated into cells with Leydig cell characteristics in vitro. Human Reproduction (Oxford, England), 34(9), 1621-1631.

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Immunohistochemical Analysis of Pancreatic Tissues

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The formalin-fixed pancreatic tissues were dehydrated and embedded in paraffin. 5-μm sections were cut. The sections were deparaffinized and then antigen-retrieved in 10 mM Tris–HCl and 1 mM EDTA (pH 8.8) for 15 min at 95 °C. The sections were incubated in PBS containing 0.1% (v/v) Triton X-100 for 5 min, then incubated in the blocking solution (PBS containing 1% [w/v] BSA and 3% [v/v] normal goat serum with or without M.O.M.™ Mouse IgG Blocking Reagent [Vector Laboratories, Burlingame, CA, USA]) for 60 min at 25 °C. The primary antibody was applied overnight at 4 °C or 25 °C, followed by incubation with a secondary antibody for 60 min at 25 °C and then with Hoechst 33258 for 10 min to stain nuclei. The treated sections were visualized with a BZ-X710 microscope (KEYENCE, Osaka, Japan) and analyzed using a BZ-X Analyzer (KEYENCE). The primary antibodies used are listed in Supplementary Table 1. The anti-Ppy antibody was prepared as reported previously [28 (link)].

Kitakaze K., Oyadomari M., Zhang J., Hamada Y., Takenouchi Y., Tsuboi K., Inagaki M., Tachikawa M., Fujitani Y., Okamoto Y, & Oyadomari S. (2021). ATF4-mediated transcriptional regulation protects against β-cell loss during endoplasmic reticulum stress in a mouse model. Molecular Metabolism, 54, 101338.

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Immunofluorescence Detection of CD45 and BCL2

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The immunofluorescence detection was performed with a Discovery XT system (Ventana Medical Systems). Tissue sections were blocked first for 30 min in Mouse IgG Blocking reagent (Vector Labs; cat. # MKB-2213) in PBS. The primary antibody incubation was performed with either mouse monoclonal Anti Human CD45 (Dako, Catalog No. M0701, 2.5 μg/ml) or rabbit polyclonal Anti BCL2 (Ventana, Catalog No. 790-4604, 0.24 μg/ml) for 6 h followed by 60 minutes incubation with a biotinylated mouse secondary antibody (Vector Labs, MOM Kit BMK-2202), at 5.75 μg/ml (1:200 dilution). The detection was performed with Secondary Antibody Blocker, Blocker D, Streptavidin-HRP D (Ventana Medical Systems), followed by incubation with Tyramide-Alexa Fluor 488 (Invitrogen, cat. #T20922).

Ramaswamy K., Forbes L., Minuesa G., Gindin T., Brown F., Kharas M.G., Krivtsov A.V., Armstrong S.A., Still E., de Stanchina E., Knoechel B., Koche R, & Kentsis A. (2018). Peptidomimetic blockade of MYB in acute myeloid leukemia. Nature Communications, 9, 110.

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Immunohistochemical Analysis of Tau Pathology

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For immunohistochemistry, we used free-floating coronal cryostat sections of the hippocampal and brainstem regions (40 μm) obtained from a wild-type, non-transgenic mouse and a Tau1N4R-G272V/P301S transgenic mouse (12 months old; Schindowski et al., 2006 (link)) and vibratome sections of the brainstem region (40 μm) obtained from a transgenic Tau0N4R-P301S mouse (Allen et al., 2002 (link)) and a transgenic Tau2N4R-P301L mouse (Terwel et al., 2005 (link)). Human biopsy samples (frontal cortex) consisted of 7 μm paraffin-embedded sections. Endogenous peroxidase activity was quenched by an H₂O₂ treatment for 30 min. Sections were either saturated by M.O.M.^TM Mouse IgG Blocking Reagent (Vector, Burlingame, CA, USA) or by normal horse serum (Vector, Burlingame, CA, USA). Overnight primary antibody incubation was done at 4°C and a biotinylated goat anti-mouse secondary antibody was used for detection. The signal was amplified using the VECTASTAIN ABC kit (Vector, Burlingame, CA, USA) and 3, 3′ diaminobenzidine tetrahydrochloride (DAB) revelation (Sigma Aldrich, Overijse, BE) was done in Tris 0.2M + H₂O₂.
The phosphorylation-specific antibody AT8 (pSer²⁰²/pThr²⁰⁵ Tau) (ThermoFisher Scientific, Waltham, MA, USA) served as reference antibody for immunohistochemistry.

Verelst J., Geukens N., Eddarkaoui S., Vliegen D., De Smidt E., Rosseels J., Franssens V., Molenberghs S., Francois C., Stoops E., Bjerke M., Engelborghs S., Laghmouchi M., Carmans S., Buée L., Vanmechelen E., Winderickx J, & Thomas D. (2020). A Novel Tau Antibody Detecting the First Amino-Terminal Insert Reveals Conformational Differences Among Tau Isoforms. Frontiers in Molecular Biosciences, 7, 48.

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