Non immune mouse igg

Manufactured by Santa Cruz Biotechnology

Non-immune mouse IgG is an immunoglobulin G (IgG) antibody derived from mouse that does not have a specific immune response. It can be used as a control in various immunological experiments.

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Lab products found in correlation

Nonimmune rabbit igg, by Santa Cruz Biotechnology (2 mentions) Protein a g sepharose beads, by Santa Cruz Biotechnology (1 mentions) Complete protease inhibitor cocktail, by Roche (1 mentions) Protein block serum, by Agilent Technologies (1 mentions) Envision polymer solution, by Agilent Technologies (1 mentions) Anti ki67 antibody, by Santa Cruz Biotechnology (1 mentions) Fluorescence microscope, by Olympus (1 mentions) Anti cd31 antibody, by Abcam (1 mentions) Protein g sepharose bead, by GE Healthcare (1 mentions) Anti rae1 antibody, by Abcam (1 mentions) Protease inhibitor cocktail, by Roche (1 mentions) Rabbit anti ve cadherin, by Cayman Chemical (1 mentions) Goat anti rabbit dylight680, by Thermo Fisher Scientific (1 mentions) Rabbit anti actin, by Merck Group (1 mentions) Rabbit anti gapdh, by Santa Cruz Biotechnology (1 mentions) Odyssey infrared imaging system, by LI COR (1 mentions) Monoclonal anti α tubulin, by Merck Group (1 mentions) Polyclonal rabbit anti nox4, by Novus Biologicals (1 mentions) Goat anti rabbit 800, by Thermo Fisher Scientific (1 mentions)

6 protocols using non immune mouse igg

Immunoprecipitation Assay Protocol

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Cells were collected and lysed in 1 ml immunoprecipitation lysis buffer (50 mM Tris-HCl [pH 7.4], 150 mM NaCl, 0.5% NP-40) with complete protease inhibitor cocktail (Roche Diagnostics). The cell lysates were precleared and then incubated with the indicated antibodies for 1 h at 4 °C. The complexes were precipitated with Protein A/G-Sepharose beads (Santa Cruz Biotechnology Inc.), washed, and resuspended in 40 μl SDS loading buffer. Non-immune mouse IgG or non-immune rabbit IgG (Santa Cruz Biotechnology Inc.) served as a negative control.

Ko H., Kim S., Yang K, & Kim K. (2018). Phosphorylation-dependent stabilization of MZF1 upregulates N-cadherin expression during protein kinase CK2-mediated epithelial-mesenchymal transition. Oncogenesis, 7(3), 27.

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Immunohistochemical Analysis of Tumor Samples

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Tumor specimens were fixed in 10 % neutral buffered formalin and embedded in paraffin. Sections were stained with hematoxylin and eosin and immunostained with anti-Ki67 antibody (Santa Cruz Biotechnology) and anti-CD31 antibody (abcam, San Francisco, CA, USA). Deparaffinized sections were pretreated by autoclaving in 10 % citric acid buffer (pH 8.0) at 120 °C for 15 min. Following treatment with protein block serum (Dako Cytomation, Kyoto, Japan) for 10 min and incubation with 2 % skim milk for 30 min to block nonspecific reactions, sections were incubated with primary antibody at 4 °C overnight. The EnVision polymer solution (horseradish peroxidase; Dako Cytomation) was then applied for 1 h. Signals were developed in 0.02 % 3,3′-diaminobenzidinetetrahydrochloride solution containing 0.1 %. As negative controls, the sections were incubated with tris(hydroxymethyl)aminomethane-buffered saline with non-immune mouse IgG (Santa Cruz Biotechnology) or non-immune rabbit IgG (Santa Cruz Biotechnology). Sections were then lightly counterstained with hematoxylin and examined under a fluorescence microscope (Olympus, Tokyo, Japan). For endothelial cell counting in tumors, five vascularized areas in a visual field (×100) were chosen, and the number of CD31⁺ cells was then counted.

Yamaguchi T., Fushida S., Yamamoto Y., Tsukada T., Kinoshita J., Oyama K., Miyashita T., Tajima H., Ninomiya I., Munesue S., Harashima A., Harada S., Yamamoto H, & Ohta T. (2015). Tumor-associated macrophages of the M2 phenotype contribute to progression in gastric cancer with peritoneal dissemination. Gastric Cancer, 19(4), 1052-1065.

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Immunoprecipitation of GABA Receptor Subunits

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Microdissected DG was obtained as described above. Lysates were obtained by passing the tissue through a 21G needle (25X) followed by agitation at room temperature (15 min) and then at 4°C (90 min). Lysates were centrifuged at 15,000 × g for 20 min to remove cell debris and pre-cleared by shaking with 40 μl of sepharose beads (1 h at 4°C). 300–350 μg of protein were incubated with 5 μg of mouse monoclonal antibodies for α₁ (NeuroMab, Davis, CA) or β_2/3 (Millipore, Billerica, MA) subunits or with 5 μg of non-immune mouse IgG (Santa Cruz Biotech, Santa Cruz, CA). After overnight incubation at 4°C, immune complexes were mixed with 25 μl of protein G-sepharose beads (GE Health Care, Piscataway, NJ) and incubated for 2 h at 4°C. Beads were washed with RIPA buffer and proteins were released in 25 μl of 2X Laemmli buffer by boiling for 3 min.

González M.I., Grabenstatter H.L., del Rio C.C., Del Angel Y.C., Carlsen J., Laoprasert R., White A.M., Huntsman M.M, & Brooks-Kayal A. (2015). Seizure-Related Regulation of GABAA Receptors in Spontaneously Epileptic Rats. Neurobiology of disease, 77, 246-256.

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ChIP-PCR Analysis of RAE1 in Breast Cancer

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ChIP analysis was performed as previously described⁴² with minor modifications. Chromatin was prepared from stable RAE1-overexpressing and control breast cancer cell lines. Briefly, 1 × 10⁶ cells were cross-linked with 1% formaldehyde for 15 min, followed by the addition of glycine at 125 mM. Chromatin was sheared by sonication to fragments averaging between 0.5 and 1 kb in buffer containing 1% SDS, 1% Triton X-100, 0.1% sodium deoxycholate, 10 mM EDTA, 50 mM Tris-HCl (pH 8.0), and protease inhibitor cocktail (Roche Applied Science, Basel, Switzerland). Chromatin was pre-cleared with protein A/G beads containing 50% slurry (Santa Cruz Biotechnology, Dallas, TX, USA) and salmon sperm DNA, followed by immunoprecipitation with anti-RAE1 antibody (Abcam) coupled to protein A/G beads under each experimental condition. Nonimmune mouse IgG (Santa Cruz Biotechnology) was used as a control. ChIP-PCR data are shown as the percentage of input after normalization with IgG. Primers for ChIP-PCR are listed in Table 2.Table 2

Primer sequences used for ChIP-PCR assay.

Amplicon sites	Sequence (5′ → 3′)
pZEB1 #1	F- GGA TCC CAC GGT TCT ACG C
pZEB1 #1	R- GCG ACC GGA GAG AGG CTA
pZEB1 #2	F- CTC ATC AAG GGA ACT CCC CG
pZEB1 #2	R- GAA TTG AGG GGC GAG GGA AA
pZEB1 #3	F- CCC ACC ACA CCT GAG GAA AA
pZEB1 #3	R- CAT GAT CCT CTC GCT TGT GTC
Gene desert	F- TGG TGG TCT GCC TTC TGC CAG T
Gene desert	R- TCA CGT GGG AGG AAG AAG TAG GGC

Oh J.H., Lee J.Y., Yu S., Cho Y., Hur S., Nam K.T, & Kim M.H. (2019). RAE1 mediated ZEB1 expression promotes epithelial–mesenchymal transition in breast cancer. Scientific Reports, 9, 2977.

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Immunoprecipitation and Western Blot Analysis of KRIT1

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Cells were treated with or without 1μM 8-pCPT-2′-O-Me-cAMP-AM (Tocris/R&D Biosystems) as indicated in the figure legends. Lysates were prepared as reported previously[12 (link)]. KRIT1 was immunoprecipitated using 2μg monoclonal anti-KRIT1 (EMD Millipore, Darmstadt, Germany). Lysates were probed with polyclonal rabbit anti-KRIT1 Δ6832 (Ginsberg Lab, UCSD) at a dilution of 1:1000. Control lysates were immunoprecipitated with mouse non-immune IgG (Santa Cruz, Dallas, TX). Antibodies used for Western blot were rabbit anti-actin (Sigma, St. Louis, MO), rabbit anti-GAPDH (Santa Cruz), rabbit anti-PLCε (Smrcka Lab, University of Rochester), and rabbit anti-VE cadherin (Cayman Chemical, Ann Arbor, MI). The secondary antibodies used were goat anti-rabbit Dylight-680 (Thermo Fisher Scientific) and goat-anti rabbit 800 (Fisher). An Odyssey Infrared Imaging System (LI-COR Biosciences, Lincoln, NE) was used to image membranes and for densitometry.

DiStefano P.V., Smrcka A.V, & Glading A.J. (2016). Phospholipase Cε Modulates Rap1 Activity and the Endothelial Barrier. PLoS ONE, 11(9), e0162338.

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Nox4 Protein Interaction Profiling

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Lysates were prepared as reported previously⁴. Nox4 was immunoprecipitated using 2 µg monoclonal anti-Nox4 antibody (Millipore/Sigma). Lysates were probed with polyclonal rabbit anti-Nox4 (Novus Biologicals, Littleton CO) at a dilution of 1:1000. Control lysates were immunoprecipitated with mouse non-immune IgG (Santa Cruz, Dallas, TX). Goat anti-rabbit Dylight-680 and goat-anti rabbit 800 (ThermoFisher) were used as secondary antibodies and the membranes imaged using an Odyssey Infrared Imaging System. NF-κB p65 was detected using a rabbit polyclonal anti-p65 (C-20) antibody from Santa Cruz. Tubulin was detected as a loading control (monoclonal anti-α-tubulin, Sigma). Goat anti-rabbit and goat anti-mouse HRP were used as a secondary antibodies and the blot was developed using Luminata™ Forte Western HRP substrate reagent, then exposed to film.

Goitre L., DiStefano P.V., Moglia A., Nobiletti N., Baldini E., Trabalzini L., Keubel J., Trapani E., Shuvaev V.V., Muzykantov V.R., Sarelius I.H., Retta S.F, & Glading A.J. (2017). Up-regulation of NADPH oxidase-mediated redox signaling contributes to the loss of barrier function in KRIT1 deficient endothelium. Scientific Reports, 7, 8296.

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