Nb100 2219

Manufactured by Novus Biologicals

Sourced in United States

The NB100-2219 is a laboratory equipment product manufactured by Novus Biologicals. It is designed for general laboratory use, but its core function is not specified in the information provided.

Automatically generated - may contain errors

Lab products found in correlation

Nb100 479, by Novus Biologicals (2 mentions) Nb100 303, by Novus Biologicals (2 mentions) Nb100 122, by Novus Biologicals (1 mentions) Anti β tubulin, by Merck Group (1 mentions) Phospho cofilin ser 3 77g2, by Cell Signaling Technology (1 mentions) Protease inhibitor cocktail, by Merck Group (1 mentions) Anti β actin, by Merck Group (1 mentions) A2066, by Merck Group (1 mentions) Sc 13119, by Santa Cruz Biotechnology (1 mentions) Sc 166553, by Santa Cruz Biotechnology (1 mentions) Nb100 310, by Novus Biologicals (1 mentions) Sc 5575, by Santa Cruz Biotechnology (1 mentions) Aebsf protease inhibitor, by Merck Group (1 mentions) Better bradford kit, by Thermo Fisher Scientific (1 mentions) Anti β actin hrp, by Merck Group (1 mentions) Nb100 449, by Novus Biologicals (1 mentions) Ecl prime western blotting detection reagent, by Cytiva (1 mentions) Lysis buffer, by Cell Signaling Technology (1 mentions) Ab5245, by Abcam (1 mentions) Pierce ecl system, by Thermo Fisher Scientific (1 mentions)

4 protocols using nb100 2219

Evaluating Hypoxia Signaling Pathways

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Lung tissue samples from mice were homogenized in RIPA lysis buffer supplemented with protease inhibitor cocktail (P8340, Sigma-Aldrich). Western blot analysis was performed using anti-PHD2 (NB100-2219 Novus Biologicals and #4835S Cell Signaling), HIF1α (NB100-479 Novus Biologicals) and HIF2α (NB100-122 Novus Biologicals) antibodies. Anti-β-actin (A2066 Sigma-Aldrich) and anti-β-tubulin antibodies (Sigma T4026) were used as a loading control. Phospho-MYPT1 (Thr696, 5163T), MYPT1 (2634T), phospho-myosin light chain 2 (Thr18/Ser19, 3674T), myosin light chain 2 (D18E2, 8505), phospho-Cofilin (Ser3, 77G2) and Cofilin (D3F9, XP® 5175T) antibodies were all from Cell Signaling Technology.

Elamaa H., Kaakinen M., Nätynki M., Szabo Z., Ronkainen V.P., Äijälä V., Mäki J.M., Kerkelä R., Myllyharju J, & Eklund L. (2022). PHD2 deletion in endothelial or arterial smooth muscle cells reveals vascular cell type-specific responses in pulmonary hypertension and fibrosis. Angiogenesis, 25(2), 259-274.

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Immunoblotting and Immunoprecipitation of HIF-1α Pathway

Cited 1 time

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Western blotting and immunoprecipitation were performed using previous methods.^{60 (link)} HIF-1α antibody (1:1 000, 10006421; Cayman), VHL antibody (1:1 000, sc-5575; Santa Cruz), PHD1 antibody (1:1 000, NB100-310; Novus), PHD2 antibody (1:1 000, NB100-2219; Novus), PHD3 antibody (1:1 000, NB100-303; Novus), HSP70 antibody (1:500, ADI-SPA-812; Enzolife), HSP90 antibody (1:500, sc-13119; Santa Cruz) and β-actin antibody (1:2 000, A2066; Sigma) were used for immunoblotting. Protein bands were quantified using ImageJ. For immunoprecipitation, nuclear extracts from cultured B cells were incubated with Dynabeads protein G and 5 μg anti-HIF-1α antibody (H1a67) overnight at 4 °C. Anti-HIF-1α, (1:1 000, 10006421; Cayman) anti-HSP70 (1:500, ADI-SPA-812; Enzolife) and anti-ubiquitin (1:500, sc-166553; Santa Cruz) antibodies were used for immunoblotting. Western blot source data are included in Fig. S9.

Meng X., Lin Z., Cao S., Janowska I., Sonomoto K., Andreev D., Katharina K., Wen J., Knaup K.X., Wiesener M.S., Krönke G., Rizzi M., Schett G, & Bozec A. (2022). Estrogen-mediated downregulation of HIF-1α signaling in B lymphocytes influences postmenopausal bone loss. Bone Research, 10, 15.

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Western Blot Analysis of HIF1α and PHDs

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Cells were lysed with 300 µL 1X Lysis buffer (Cell Signaling Technologies, Inc., Danvers, MA, USA) with 1 μg·mL⁻¹ 4-(2-aminoethyl) benzenesulfonyl fluoride hydrochloride (AEBSF) protease inhibitor (Sigma-Aldrich, Inc.). Total cell lysate protein concentrations were determined with the Better Bradford Kit (Thermo-Fisher Scientific) according to the manufacturer’s instructions. Western blot analysis was performed with 50 µg of MPC2 or 40 µg of U2OS cellular lysates. The blots were incubated with primary antibody to hypoxia-inducible factor 1-alpha (HIF1α) (NB100–449; Novus Biologicals, Littleton, CO, USA) Phd1 (Abcam; ab108980), Phd2 (Novus; NB100-2219), Phd3 (Novus: NB100-303) overnight, then incubated with secondary antibody at 1:2 000 (anti-rabbit–horseradish peroxidase [HRP]; Cell Signaling Technologies). Blots were stripped using SDS-glycine and reprobed with 1:15 000 anti-β-actin-HRP (A3854; Sigma-Aldrich). Detection was performed using the ECL Prime Western Blotting Detection Reagents (Amersham-GE Healthcare, Pittsburgh, PA, USA) and the GE AB1600 digital imager.

Noonan M.L., Ni P., Solis E., Marambio Y.G., Agoro R., Chu X., Wang Y., Gao H., Xuei X., Clinkenbeard E.L., Jiang G., Liu S., Stegen S., Carmeliet G., Thompson W.R., Liu Y., Wan J, & White K.E. (2023). Osteocyte Egln1/Phd2 links oxygen sensing and biomineralization via FGF23. Bone Research, 11, 7.

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Cardiac Protein Detection and Quantification

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For GATA4, pGATA4 and BNP thirty μg and for MEF2C hundred μg of total protein from heart tissue was lysed in EBC + urea buffer (50 mM Tris-HCl pH 7.5, 150 mM NaCl, 0.5% NP-40, 3 M urea) resolved by SDS-PAGE, transferred to PVDF membrane, blotted, and probed with the following primary antibodies: anti-GATA4 pSer105 (1:600 ab5245, Abcam), anti-GATA4 (1:1000 sc-9053, Santa Cruz Biotechnology), anti-BNP (1:500 ab236101, Abcam), anti-vinculin (1:200 V4505, Merck) and anti-MEF2C (1:7500 sc-313×, Santa Cruz Biotechnology) followed by a HRP-conjugated secondary antibody (1:5000; Bio-Rad). For blotting of HIF1α and HIF-P4H-2 hundred ug of total protein from heart tissue lysed using EBC + urea lysis buffer and was resolved by SDS-PAGE, transferred with nitrocellulose membrane, blotted and probed with anti-HIF1α (1:500, NB100-479 Novus Biologicals) and anti-HIF-P4H-2 (1:500, Novus Biologicals NB100-2219)primary antibodies followed by HRP-conjugated secondary antibody. The Pierce ECL system (Ther-moScientific) was used for detection. Western blot images were quantified with fiji (ImageJ) software. For original blots see (Supplementary Fig. S1-S8).

Röning T., Magga J., Laitakari A., Halmetoja R., Tapio J., Dimova E.Y., Szabo Z., Rahtu-Korpela L., Kemppi A., Walkinshaw G., Myllyharju J., Kerkelä R., Koivunen P, & Serpi R. (2022). Activation of the hypoxia response pathway protects against age-induced cardiac hypertrophy. Journal of molecular and cellular cardiology, 164.

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