Rabbit monoclonal antibodies against

Manufactured by Cell Signaling Technology

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Rabbit monoclonal antibodies are a type of laboratory reagent used in various research applications. They are produced by immunizing rabbits with a specific antigen, and then isolating and purifying the antibodies that are generated in response. These antibodies can be used to detect and quantify the presence of target proteins or other biomolecules in biological samples, such as cell lysates or tissue extracts.

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Lab products found in correlation

Human α thrombin, by Enzyme Research (3 mentions) P erk1 2, by Cell Signaling Technology (2 mentions) Erk1 2, by Cell Signaling Technology (2 mentions) 4 6 diamidino 2 phenylindole (dapi), by Thermo Fisher Scientific (2 mentions) Rabbit monoclonal antibodies against, by Abcam (1 mentions) β actin, by Cell Signaling Technology (1 mentions) Horseradish peroxidase conjugated secondary antibody, by Novus Biologicals (1 mentions) Imagequanttm las 500, by GE Healthcare (1 mentions) Apoptozole, by MedChemExpress (1 mentions) C jun n terminal kinase jnk, by Cell Signaling Technology (1 mentions) P jnk, by Cell Signaling Technology (1 mentions) 3 methyladenine 3 ma, by Santa Cruz Biotechnology (1 mentions) Rabbit polyclonal antibodies against β actin, by Abcam (1 mentions) Cisplatin, by Merck Group (1 mentions) Fenofibrate, by Merck Group (1 mentions) Ko143, by Enzo Life Sciences (1 mentions) Trypsin, by Corning (1 mentions) Hrp labeled goat anti rabbit secondary antibody, by Cell Signaling Technology (1 mentions) Geneticin g418, by Enzo Life Sciences (1 mentions) Dulbecco s modified eagle s medium dmem, by Corning (1 mentions)

12 protocols using rabbit monoclonal antibodies against

Quercetin Modulates Inflammatory Pathways in LPS-Stimulated Cells

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Quercetin was bought from the National Institute for Pharmaceutical and Biological Products Control (Beijing, China). 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide (MTT) and bicinchoninic acid (BCA) protein concentration determination kits were purchased from Solarbio Life Science Co., Ltd (Beijing, China). LPS from P. gingivalis was supplied by Invitrogen (St. Louis, CA, USA). Rabbit monoclonal antibodies against IκBα, phosphorylated IκBα (p-IκBα), p65 subunit of nuclear factor-kappa B (NF-κB) (p65), phosphorylated p65 (p-p65) and β-actin were purchased from Cell Signaling Technology Inc (Beverly, MA, USA), with Rabbit monoclonal antibodies against PPAR-γ, LXRα and TLR4 from Abcam Inc. (Cambridge, UK). Horseradish peroxidase (HRP)-conjugated goat anti-rabbit IgG was provided by Biosynthesis Biotechnolgogy Co., Ltd (Beijing,China). Enhanced chemiluminescence (ECL) kit was bought from Yanxi Co., Ltd (Shanghai, China). And the enzyme-linked immunosorbent assay (ELISA) kits were provided by Westtang Co., Ltd (Shanghai, China). All the other reagents used in the present work are of analytical grade.

Xiong G., Ji W., Wang F., Zhang F., Xue P., Cheng M., Sun Y., Wang X, & Zhang T. (2019). Quercetin Inhibits Inflammatory Response Induced by LPS from Porphyromonas gingivalis in Human Gingival Fibroblasts via Suppressing NF-κB Signaling Pathway. BioMed Research International, 2019, 6282635.

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Western Blot Analysis of STAT3 and P-STAT3

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Total proteins were extracted from frozen tumors using RIPA buffer supplemented with protease inhibitor. Then, proteins were resolved on a SDS-PAGE and moved to polyvinylidene difluoride (PVDF) membrane [87 ]. Blocking of the membranes was achieved by using 3% bovine serum albumin in Tris-buffered saline with Tween 20 (TBST80) 60 min at room temperature. Next, membranes were incubated in primary mouse specific rabbit monoclonal antibodies against STAT3 (#4904, 1:2000), rabbit monoclonal antibodies against P-STAT3 (#9145, dilution 1:2000), and mouse monoclonal antibodies against β-actin (#3700, dilution 1:1000) from Cell signaling technology, Danvers, MA, USA) overnight at 4 °C. Then, membranes were incubated with horseradish peroxidase-conjugated secondary antibody (Novus Biologicals, Centennial, CO, USA). Blots were detected with a chemiluminescent kit (Bio-Rad catalog #170-5060) and imaged (ImageQuantTMLAS500, GE Healthcare Life Sciences). The intensity of target proteins was normalized to the β-actin protein using Image-J1.52p (NIH, Bethesda, MD, USA).

El-Sherbiny M., El-Sayed R.M., Helal M.A., Ibrahiem A.T., Elmahdi H.S., Eladl M.A., Bilay S.E., Alshahrani A.M., Tawfik M.K., Hamed Z.E., Mohamed A.O, & Zaitone S.A. (2021). Nifuroxazide Mitigates Angiogenesis in Ehlrich’s Solid Carcinoma: Molecular Docking, Bioinformatic and Experimental Studies on Inhibition of Il-6/Jak2/Stat3 Signaling. Molecules, 26(22), 6858.

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Modulating HSP70 Activity: Protocols

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TRC051384 (termed as heat shock protein 70(HSP70) agonist, catalog no. HY-101712) and apoptozole (termed as HSP70 inhibitor; catalog no. HY-15098) were purchased from MedChemExpress (New Jersey, USA). Rabbit monoclonal antibodies against PCSK9 (catalog no. 85813) for western blotting, HSP70(catalog no. 4873), p38 (catalog no. 8690), phosphorylated (p)−p38(catalog no. 9215), extracellular signal−regulated kinase 1/2 (ERK1/2; catalog no. 4695), p−ERK1/2 (catalog no. 4370), c−Jun N−terminal kinase (JNK; catalog no. 9252), and p−JNK (catalog no. 9251) were acquired from Cell Signaling Technology (Danvers, MA, USA). Rabbit monoclonal antibody against PCSK9 (catalog no. ab28770) for immunohistochemistry, immunoprecipitation and immunofluorescence was purchased from Abcam (Cambridge, MA, USA). Mouse monoclonal antibody against β−actin (AA128) was purchased from Biotime Biotechnology Co. Ltd. (Shanghai, China).

Xu B., Li S., Fang Y., Zou Y., Song D., Zhang S, & Cai Y. (2021). Proprotein Convertase Subtilisin/Kexin Type 9 Promotes Gastric Cancer Metastasis and Suppresses Apoptosis by Facilitating MAPK Signaling Pathway Through HSP70 Up-Regulation. Frontiers in Oncology, 10, 609663.

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Fenofibrate and Cisplatin Apoptosis Pathway

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Fenofibrate and cisplatin (SIGMA-Aldrich, St. Louis, MO, USA) were used. Rabbit monoclonal antibodies against human p53 upregulated modulator of apoptosis (Puma), mouse cleaved caspase-8, human p38 and human NFkB, phosphorylated human JNK (p-JNK), human p38 (p-p38), human ERK (p-ERK), human NFkB (p-NFkB) and human 14-3-3 (p-14-3-3), and rabbit polyclonal antibodies against human cleaved caspase-3, human JNK, rat ERK, human 14-3-3, mouse cleaved caspase-9, mouse caspase 12, human AMPK and human LC3 were purchased from Cell Signaling Technology (Boston, USA). Rabbit polyclonal antibody against human p62 was purchased from MEDICAL and BIOLOGICAL LABORATORIES Co. Ltd. (Nagano, Japan). Rabbit polyclonal antibodies against β-actin, and Rabbit monoclonal antibodies against human Cox IV were purchased from Abcam Inc. (Cambridge, UK). Horseradish peroxidase (HRP)–conjugated anti-mouse or anti-rabbit immunoglobulins (Dako, Glostrup, Denmark) were also used. An inhibitor of autophagy, 3-methyladenine (3-MA), was purchased from Santa Cruz Biotechnology (Texas, U.S.A).

Kimura H., Kamiyama K., Imamoto T., Takeda I., Masunaga S., Kobayashi M., Mikami D., Takahashi N., Kasuno K., Sugaya T, & Iwano M. (2022). Fenofibrate reduces cisplatin-induced apoptosis by inhibiting the p53/Puma/Caspase-9 pathway and the MAPK/Caspase-8 pathway rather than by promoting autophagy in murine renal proximal tubular cells. Biochemistry and Biophysics Reports, 30, 101237.

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Topotecan Resistance Mechanism Study

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Topotecan (TPT) was purchased from Chemitek (Indianapolis, IN, United States). The mouse monoclonal anti-P-glycoprotein antibody, and 3-(4, 5-dimethylthiazol-yl)-2, 5-diphenyltetrazolium bromide (MTT), and dimethyl sulfoxide (DMSO), were supplied from MilliporeSigma Co. (St. Louis, MO, United States). Dulbecco’s modified Eagle’s medium (DMEM), 0.25% trypsin, and fetal bovine serum (FBS) were purchased from Corning Inc. (New York, NY, United States). The rabbit monoclonal antibodies against human MRP1/ABCC1 and human DNA topoisomerase I, the HRP-labeled goat anti-rabbit secondary antibody, and the HRP-linked rabbit anti-mouse secondary antibody were supplied from Cell Signaling (Danvers, MA, United States). Ko143, mitoxantrone (MX), cisplatin, and geneticin (G418) were obtained from Enzo life Sciences (Farmingdale, NY, United States). The mouse monoclonal antibodies for ABCG2 and glyceraldehyde phosphate dehydrogenase (GAPDH), the Alexa Fluor 488-labeled rabbit anti-mouse secondary antibody, 4,6-diamidino-2-phenylindole (DAPI), and all other chemicals were obtained from Thermo Fisher Scientific Inc. (Rockford, IL, United States).

Lei Z.N., Teng Q.X., Zhang W., Fan Y.F., Wang J.Q., Cai C.Y., Lu K.W., Yang D.H., Wurpel J.N, & Chen Z.S. (2020). Establishment and Characterization of a Topotecan Resistant Non-small Cell Lung Cancer NCI-H460/TPT10 Cell Line. Frontiers in Cell and Developmental Biology, 8, 607275.

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OSCC Cell Line Signaling Pathway

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OSCC cell lines treated with vehicle, Gefitinib (500 nM) and/or TAE684 (500 nM) for 6 h were harvested and lysed in 1% Triton-PBS. Cell lysates (100units/A280) were used for protein electrophoresis in 10% SDS–PAGE. SDS–PAGE separated proteins were transferred to PVDF membrane and the membrane blocked in 5% milk. Rabbit monoclonal antibodies against STAT3 (1:2000; #12640), p-STAT3(Tyr705) (1:4000; #9145) and ERK1/2 (1:2000; #4695) were used (Cell Signaling; Danvers, MA). Rabbit polyclonal antibodies against AKT (1:2000; #9272), p-AKT(Ser473) (1:4000; #9271), EGFR (1:1000, #4267), p-EGFR(Y1148) (1:4000; #4404) and p-ERK1/2 (1:5000; #9101) were also used (Cell Signaling; Danvers, MA). GAPDH rabbit polyclonal antibodies (1:1000; Rockland, #600-401-A33, Limerick, PA) were used to confirm equal loading of protein. Primary antibodies were diluted in a total of 5 ml diluent (1% milk in PBS-0.1% Tw-20) and the membrane incubated overnight at 4 °C. The membrane was washed 3× with PBS-Tw-20, incubated with ECL Plus detection solution (GE Healthcare, South San Francisco, CA) for 1 min and signal detected by exposure to radiograph film for 30 sec. Band intensities were quantified using ImageJ software.

Gonzales C.B., De La Chapa J.J., Saikumar P., Singha P.K., Dybdal-Hargreaves N.F., Chavez J., Horning A.M., Parra J, & Kirma N.B. (2016). Co-targeting ALK and EGFR parallel signaling in oral squamous cell carcinoma. Oral oncology, 59, 12-19.

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Immunoblotting of NF-κB Pathway

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RD was purchased from Guangzhou Honsea Industry Co. Ltd. Rabbit monoclonal antibodies against phospho-Ser32 IκBα (#2859), p65 (#8242), phospho-Ser536 p65 (#3033), β-actin (#4970) and a mouse monoclonal antibody against IκBα (#4814) were purchased from Cell Signaling Technology (Cell Signaling, USA). Goat polyclonal secondary antibodies against mouse (#A0216) or rabbit (#A0208) were purchased from Beyotime Biotechnology (Beyotime, China).

Hu Q., Lu Y., Hu F., He S., Xu X., Niu Y., Zhang H., Li X, & Su Q. (2020). Resistant dextrin reduces obesity and attenuates adipose tissue inflammation in high-fat diet-fed mice. International Journal of Medical Sciences, 17(17), 2611-2621.

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Molecular Mechanisms of SLC41A1 in Apoptosis Regulation

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Crystal violet, the caspase inhibitor Z-VAD-FMK, and the Akt activator SC79 were purchased from Sigma-Aldrich (St. Louis, MO). JC-1 dye was purchased from Life Technologies (Carlsbad, CA). A non-viral plasmid expressing SLC41A1 open reading frame (ORF) was obtained from Origene Technologies (Rockville, MD). Scrambled negative control small interfering RNA (siRNA) and siRNA against Bcl-2-associated X protein (Bax) were purchased from Santa Cruz Biotechnology (Dallas, TX). Rabbit polyclonal antibody against SLC41A1 (#ab83701) was obtained from Abcam (Cambridge, UK). Rabbit monoclonal antibodies against Akt (#4685), phospho-Akt (p-Akt, Ser473, #4060), mechanistic target of rapamycin (mTOR, #2983), p-mTOR (Ser2448, #5536), Bax (#5023), cytochrome c (#4280), cleaved caspase-3 (#9664), and GAPDH (#5174) were purchased from Cell Signaling Technology (Danvers, MA).

Xie J., Cheng C.S., Zhu X.Y., Shen Y.H., Song L.B., Chen H., Chen Z., Liu L.M, & Meng Z.Q. (2019). Magnesium transporter protein solute carrier family 41 member 1 suppresses human pancreatic ductal adenocarcinoma through magnesium-dependent Akt/mTOR inhibition and bax-associated mitochondrial apoptosis. Aging (Albany NY), 11(9), 2681-2698.

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Platelet Activation Signaling Study

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Luciferin/luciferase reagent and collagen were purchased from Chronolog, Havertown, PA. Human α-thrombin was from Enzyme Research Laboratories, South Bend, IN. CD40L was purchased from eBioscience (San Diego, CA, USA). Rabbit monoclonal antibodies against phosphorylated Ser⁴⁷³ residue of Akt and against phosphorylated Thr¹⁸⁰/Tyr¹⁸² residues of p38 MAPK were from Cell Signaling Technology (Beverly, MA, USA). A rabbit polyclonal antibody against TRAF3 was purchased from Santa Cruz Biotechnology Inc. (Santa Cruz, CA, USA). A rabbit polyclonal antibody against TRAF2 was purchased from NeoBiolab (Woburn, MA, USA). Fluorescein isothiocyanate (FITC)-conjugated rat anti-mouse P-selectin and integrin β3 antibodies were from BD Pharmingen.

Zhang R., Zhang G., Xiang B., Chen X., Tang L., Shi S., Liu Y., Ai X., Xie P, & Li Z. (2017). TRAF3 negatively regulates platelet activation and thrombosis. Scientific Reports, 7, 17112.

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Quantifying NF-κB p65 Nuclear Localization

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Cells were seeded on glass coverslips, fixed with methanol for 15 min at −20 °C, and blocked with Block-Ace™ (KAC Co., Ltd.) for 1 h at room temperature. The cells were then incubated with rabbit monoclonal antibodies against NF-κB p65 (Cat#8242, Cell Signaling Technology) for 1 h at room temperature, followed by an Alexa Fluor 488-conjugated goat-anti-rabbit secondary antibody (Thermo Fisher Scientific, Waltham, MA, USA) for 1 h at room temperature. The signals were detected, and images were acquired using an All-in-One Fluorescence Microscope BZ-X810 (KEYENCE, Osaka, Japan). The images were taken randomly at 5 points per glass coverslip in each sample to calculate the NF-κB p65 nuclear localization proportion. The cells were considered positive for nuclear localization of NF-κB p65 if the fluorescence intensity of their nuclei exceeded that of their cytoplasm. In addition, we calculated the percentage of positive for nuclear localization of NF-κB p65 to total cells in these points for each sample.

Tanabe N., Tomita K., Manaka S., Ichikawa R., Takayama T., Kawato T., Ono M., Masai Y., Utsu A., Suzuki N, & Sato S. (2023). Co-Stimulation of AGEs and LPS Induces Inflammatory Mediators through PLCγ1/JNK/NF-κB Pathway in MC3T3-E1 Cells. Cells, 12(10), 1383.

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