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13c glutamine

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13C-glutamine is a stable isotope-labeled compound used in research and analytical applications. It contains a carbon-13 label, which allows for the tracking and quantification of glutamine metabolism in biological systems. This product serves as a versatile tool for researchers studying cellular biochemistry and metabolism.

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Lab products found in correlation

Rpmi 1640 medium, by Thermo Fisher Scientific (2 mentions) 7890b 5977b gc ms system, by Agilent Technologies (1 mentions) Masshunter quantitative analysis software, by Agilent Technologies (1 mentions) Db 5ms column, by Agilent Technologies (1 mentions) Uplc hss t3 column, by Thermo Fisher Scientific (1 mentions) 1290 uhplc system, by Agilent Technologies (1 mentions) Q exactive mass spectrometer, by Thermo Fisher Scientific (1 mentions) Asparaginase, by Merck Group (1 mentions) Glutamine, by Merck Group (1 mentions) Anti pd 1 antibody pembrolizumab, by Selleck Chemicals (1 mentions) Nk 92, by American Type Culture Collection (1 mentions)

4 protocols using 13c glutamine

1

Quantifying Cellular Glutamine Uptake

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Cells were washed with glutamine-free RPMI-1640 medium (Thermo Fisher, MA, USA) and incubated with 4 mmol/L 13C-glutamine (Sigma–Aldrich, MO, USA) in RPMI-1640 medium for 12 h. Then, the lysates were evaluated to determine the [13C] level using a scintillation counter to analyse relative glutamine uptake. Radioactivity was measured as counts per minute (CPM) and normalized to the protein concentration.
Shi J., Ju R., Gao H., Huang Y., Guo L, & Zhang D. (2021). Targeting glutamine utilization to block metabolic adaptation of tumor cells under the stress of carboxyamidotriazole-induced nutrients unavailability. Acta Pharmaceutica Sinica. B, 12(2), 759-773.
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2

Isotope Tracing of Cellular Metabolism

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RPE media were replaced with DMEM supplemented with FBS (1%), glucose (5.5mM) and 13C glutamine (200μM, Sigma) or 13C proline (200μM, Sigma) for 24 hours. Metabolites were extracted in 80% cold methanol, dried, derivatized by methoxyamine and N-tertbutyldimethylsilyl-N-methyltrifluoroacetamide, and analyzed by an Agilent 7890B/5977B GC/MS system with an Agilent DB-5MS column (30 m × 0.25 mm × 0.25 μm film). Mass spectra were collected from m/z 80–600 under selective ion monitoring mode. The data was analyzed by Agilent MassHunter Quantitative Analysis Software.
Engel A.L., Wang Y., Khuu T.H., Worrall E., Manson M.A., Lim R.R., Knight K., Yanagida A., Qi J.H., Ramakrishnan A., Weleber R.G., Klein M.L., Wilson D.J., Anand-Apte B., Hurley J.B., Du J, & Chao J.R. (2021). Extracellular Matrix Dysfunction in Sorsby Patient-Derived Retinal Pigment Epithelium. Experimental eye research, 215, 108899.
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3

Quantifying Glutamine Metabolism

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To assess the glutamine uptake and glutamine-derived metabolites, cells were cultured in glutamine-free medium supplemented with 2 mM 13C-glutamine (Sigma-Aldrich). After incubation for 24 hours, cell lysis was collected for metabolic analysis using an UHPLC system (1290, Agilent Technologies) with a UPLC HSS T3 column (2.1 mm×100 mm, 1.8μm) coupled to Q Exactive mass spectrometer (Thermo). The raw data were converted to the mzXML format using ProteoWizard and processed with an in-house program, which was developed using R and based on XCMS [23] (link), for peak detection, extraction, alignment, and integration. In-house MS2 database (BiotreeDB, Shanghai, China) was applied in metabolite annotation.
Xiong J., Wang N., Zhong H.J., Cui B.W., Cheng S., Sun R., Chen J.Y., Xu P.P., Cai G., Wang L., Sun X.J., Huang J.Y, & Zhao W.L. (2021). SLC1A1 mediated glutamine addiction and contributed to natural killer T-cell lymphoma progression with immunotherapeutic potential. EBioMedicine, 72, 103614.
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4

Culturing and Characterizing NK Cell Lines

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Natural-killer cell line NK-92 (Cat# CRL-2407) was available from American Type Culture Collection (Manassas, VA, USA). SNK-6 was kindly provided by Professor Norio Shimizu and Yu Zhang of Chiba University. Cell lines were authenticated using Short Tandem Repeat (STR) analysis (Genetic Testing Biotechnology, STR Profile Reports in Supplemental Data). Recent mycoplasma testing has been performed with Lonza LT07-705 MycoAlert™ PLUS Mycoplasma Detection Kit. NK-92 cells were cultured in α-MEM medium supplemented with 10% FBS, 10% HBS and recombinant human IL-2 (20ng/ml). SNK-6 cells were cultured in RPMI-1640 medium supplemented with 10% FBS, 10% HBS and recombinant human IL-2 (80ng/ml). Asparaginase (Cat# A3809), 13C-glutamine (Cat# 184161-19-1), and glutamine (Cat# 56-85-9) were obtained from Sigma-Aldrich (St.Louis, MO, USA). Anti-PD-1 antibody pembrolizumab (Cat# A2005) was from Selleck Chemicals (Houston, TX, USA).
Xiong J., Wang N., Zhong H.J., Cui B.W., Cheng S., Sun R., Chen J.Y., Xu P.P., Cai G., Wang L., Sun X.J., Huang J.Y, & Zhao W.L. (2021). SLC1A1 mediated glutamine addiction and contributed to natural killer T-cell lymphoma progression with immunotherapeutic potential. EBioMedicine, 72, 103614.
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