Sc 293121

Manufactured by Santa Cruz Biotechnology

Sourced in United States

Sc-293121 is a laboratory equipment product offered by Santa Cruz Biotechnology. It is a device designed for scientific research purposes. The core function of Sc-293121 is to facilitate specific laboratory processes, but a detailed description cannot be provided while maintaining an unbiased and factual approach.

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Anti-NANOG (sc-293121, (2 citations)

4 protocols using sc 293121

Characterizing Pluripotent Stem Cells

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To detect targeted antigens and p53 in pluripotent stem cells, we immobilized cells with PBS containing 4% polyformaldehyde at room temperature for 10 minutes. After washing with PBS, the cells were incubated in PBS containing 0.1% Triton X-100 for 20 minutes at room temperature. Then, we stained fixed cells with SSEA-4 (1:100; monoclonal antibody; MAB8490; Stemgent), TRA-1-60 (1/200; monoclonal antibody; 09–0010; Stemgent), OCT4 (1/200; monoclonal antibody; MAB4419A4; Millipore), Nanog (1/600; monoclonal antibody; sc-293121; Santa Cruz) and p53 (1/500; monoclonal antibody; ab1101; abcam). These primary antibodies were visualized by with goat anti-rabbit IgG bound to Alexa 488 and goat anti-rabbit IgG bound to Alexa 594 or goat anti-mouse IgG bound to Alexa Fluor 488. Nuclear staining was performed with DAPI, and fluorescence images were obtained using Zeiss inverted LSM confocal microscopy (Carl Zeiss).

Li Y., Li T., Tang Y., Zhan Z., Ding L., Song L., Yu T., Yang Y., Ma J., Zhang Y., Zhou Y., Gu S., Xu M., Gao Y, & Li Y. (2020). The function of a heterozygous p53 mutation in a Li-Fraumeni syndrome patient. PLoS ONE, 15(6), e0234262.

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Western Blot Analysis of Stem Cell Markers

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Cell lysates and protein samples were prepared as previously reported [60 (link)]. Proteins were resolved by SDS-polyacrylamide gel electrophoresis and transferred to a nitrocellulose membrane (Bio-Rad) for detection with primary antibodies against SPIN1 (diluted 1:500, 19531-1-AP; Proteintech Group, Inc., Rosemont, IL, USA), CTNNB1 (diluted 1:500, sc-59737, Santa Cruz Biotechnology), MYC (diluted 1:500, sc-40, Santa Cruz Biotechnology), PTEN (diluted 1:500, sc-9145, Santa Cruz Biotechnology), AKT1/2/3 (diluted 1:500, sc-8312, Santa Cruz Biotechnology), p-AKT1/2/3 (Ser 473)-R (diluted 1:500, sc-7985-R, Santa Cruz Biotechnology), OCT 3/4 (diluted 1:300, sc-5279, Santa Cruz Biotechnology), SOX2 (diluted 1:300, sc-20088, Santa Cruz Biotechnology), NANOG (diluted 1:300, sc-293121, Santa Cruz Biotechnology) and ACTIN (diluted 1:500, A5060, Sigma-Aldrich). The membranes were then incubated with the appropriate horseradish peroxidase (HRP)-conjugated secondary antibodies (diluted 1:5000, Perce, Thermo Fisher Scientific). The protein bands were revealed with an enhanced chemiluminescence detection system (ECL; Bio-Rad) and visualized by ChemiDoc XRS system (Bio-Rad) and Quality One 4.5.2 (Biorad) software. Finally, protein levels were normalized using ACTIN levels. Protein bands were quantified densitometrically.

Drago-Ferrante R., Pentimalli F., Carlisi D., Blasio A.D., Saliba C., Baldacchino S., Degaetano J., Debono J., Caruana-Dingli G., Grech G., Scerri C., Tesoriere G., Giordano A., Vento R, & Fiore R.D. (2017). Suppressive role exerted by microRNA-29b-1-5p in triple negative breast cancer through SPIN1 regulation. Oncotarget, 8(17), 28939-28958.

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Protein Expression Analysis of A549 Cells

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Collected A549 and A549/GR cells were lysed in RIPA buffer (150 mM NaCl, 1% Nonidet p-40, 50 mM Tris, pH 8.0, and a protease inhibitor cocktail). Thirty micrograms of each sample was separated by SDS-PAGE and transferred to a nitrocellulose membrane (Bio-Rad, USA). Western blotting was performed with rabbit or mouse antibodies against Prx II: LF-PA0091 (AbFrontier, Seoul, South Korea), CD133: PA2049 (Boster Bio, CA, USA), Nanog: sc-293121, OCT3/4: sc-9081, Sox2: sc-365823 (Santa Cruz Biotechnology), VEGFR2: sc-6251, E-cad: sc-7870, Vimentin: sc-6260 (Santa Cruz Biotechnology), Shh: 2207 s, Gli-1: 3538 s, Notch 1: 3268 s (Cell Signaling Technology), CXCR4: YF-MA16239, STAT3: LF-MA30485,pSTAT3 (Tyr 705): LF-PA20474, pSTAT3 (Tyr-727): LF-PA20473 Hes-1: YF-MA11051, and β-Catenin: YF-MA10213 (AbFrontier). Protein expression levels were detected using Super signal West Pico PLUS Chemiluminescent Substrate (Thermo Fisher, #34577).

Chandimali N., Huynh D.L., Zhang J.J., Lee J.C., Yu D.Y., Jeong D.K, & Kwon T. (2018). MicroRNA-122 negatively associates with peroxiredoxin-II expression in human gefitinib-resistant lung cancer stem cells. Cancer Gene Therapy, 26(9), 292-304.

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Immunofluorescent Staining of Pluripotent Stem Cells

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iPS cells were grown on 15 mm glass coverslips coated with iMatrix-511. Cells were fixed with 4% paraformaldehyde diluted in phosphate-buffered saline (PBS) for 15 min at room temperature, permeabilized with 0.1% Triton X-100 diluted in PBS and blocked with blocking buffer (0.1% Tween-20 and 1% bovine serum antigen diluted in PBS). Then, cells were stained with anti-OCT4 (ab19857, Abcam) and anti-NANOG (clone 1E6C4, sc-293121, Santa Cruz) diluted 500-fold in blocking buffer for 1 h at room temperature. The cells were then rinsed three times for 5 min each with 0.1% Tween-20 in PBS. Subsequently, the cells were stained with Alexa Fluor 488-conjugated goat anti-rabbit IgG (A-11008, Invitrogen) and Alexa Fluor 568-conjugated goat anti-mouse IgG (A-11004, Invitrogen) diluted 1000-fold in blocking buffer for 1 h at room temperature and then rinsed three times for 5 min each with 0.1% Tween-20 in PBS. Finally, cells were stained with 2 µg/μl DAPI (Dojindo, D523) in PBS for 5 min at room temperature. Images were acquired with an LSM780 confocal microscope (Zeiss) and Zen Software Black Edition (Zeiss).

Ohno T., Akase T., Kono S., Kurasawa H., Takashima T., Kaneko S, & Aizawa Y. (2022). Biallelic and gene-wide genomic substitution for endogenous intron and retroelement mutagenesis in human cells. Nature Communications, 13, 4219.

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