2 mercaptoehanol, by Thermo Fisher Scientific - Product details

1

PDAC Organoids Potentiate CD8+ T Cell Killing

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Mouse PDAC cells stably expressing OVA and carrying doxycycline (Dox)-inducible mTurquoise2-Atg4B^C74A were grown as organoids, treated with or without Dox (1 μg/mL) for 96 hrs. Organoids were dissociated into single cells, which were then incubated with either anti-H-2K^b-SIINFEKL antibody (clone 25-D1.16, BioXCell, BE0207) or isotype control (clone MOPC-21, BioXCell, BE0083) at 100 μg/mL for 30 min at 4 °C. Total splenocytes were harvested from OT-I mice and CD8⁺ T cells were enriched using Dynabeads® Untouched™ Mouse CD8 Cells (Invitrogen, 11417D) following manufacturer’s instructions. Isolated CD8⁺ T cells were labelled with 10 μM CFSE (BioLegend) for 10 min at RT in the dark, washed 3x with RPMI-1640 supplemented with 10% FBS. Ten thousand PDAC cells and forty thousand CD8⁺ cells were seeded in 96-well plates and cultured in 100 μL 50% DMEM and 50% RPMI-1640 supplemented with 10% FBS, 10 ng/mL recombinant murine IL-2 (Peprotech), 27.5 μM 2-Mercaptoehanol (Gibco), and 100 μg/mL of respective antibodies. After 48 hrs, CD8⁺ T cells were harvested and stained with anti-CD8a antibody (AF647, clone 53–6.7, BioLegend) and DAPI, and proliferation was analyzed by CFSE dilution using flow cytometry. After removal of CD8⁺ T cells, the viability of remaining PDAC cells were measured by CellTiter-Glo (Promega).

Yamamoto K., Venida A., Yano J., Biancur D.E., Kakiuchi M., Gupta S., Sohn A.S., Mukhopadhyay S., Lin E.Y., Parker S.J., Banh R.S., Paulo J.A., Wen K.W., Debnath J., Kim G.E., Mancias J.D., Fearon D.T., Perera R.M, & Kimmelman A.C. (2020). Autophagy promotes immune evasion of pancreatic cancer by degrading MHC-I. Nature, 581(7806), 100-105.

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PDAC Organoids Potentiate CD8+ T Cell Killing

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Mouse PDAC cells stably expressing OVA and carrying doxycycline (Dox)-inducible mTurquoise2-Atg4B^C74A were grown as organoids, treated with or without Dox (1 μg/mL) for 96 hrs. Organoids were dissociated into single cells, which were then incubated with either anti-H-2K^b-SIINFEKL antibody (clone 25-D1.16, BioXCell, BE0207) or isotype control (clone MOPC-21, BioXCell, BE0083) at 100 μg/mL for 30 min at 4 °C. Total splenocytes were harvested from OT-I mice and CD8⁺ T cells were enriched using Dynabeads® Untouched™ Mouse CD8 Cells (Invitrogen, 11417D) following manufacturer’s instructions. Isolated CD8⁺ T cells were labelled with 10 μM CFSE (BioLegend) for 10 min at RT in the dark, washed 3x with RPMI-1640 supplemented with 10% FBS. Ten thousand PDAC cells and forty thousand CD8⁺ cells were seeded in 96-well plates and cultured in 100 μL 50% DMEM and 50% RPMI-1640 supplemented with 10% FBS, 10 ng/mL recombinant murine IL-2 (Peprotech), 27.5 μM 2-Mercaptoehanol (Gibco), and 100 μg/mL of respective antibodies. After 48 hrs, CD8⁺ T cells were harvested and stained with anti-CD8a antibody (AF647, clone 53–6.7, BioLegend) and DAPI, and proliferation was analyzed by CFSE dilution using flow cytometry. After removal of CD8⁺ T cells, the viability of remaining PDAC cells were measured by CellTiter-Glo (Promega).

Yamamoto K., Venida A., Yano J., Biancur D.E., Kakiuchi M., Gupta S., Sohn A.S., Mukhopadhyay S., Lin E.Y., Parker S.J., Banh R.S., Paulo J.A., Wen K.W., Debnath J., Kim G.E., Mancias J.D., Fearon D.T., Perera R.M, & Kimmelman A.C. (2020). Autophagy promotes immune evasion of pancreatic cancer by degrading MHC-I. Nature, 581(7806), 100-105.

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Synthesis and Characterization of Hydrogel Polymers

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N-isopropylacrylamide (NIPAAm, cat. no. 415324), N-(hydroxymethyl) acrylamide, (NHMAAm, cat. no. 245801), N,N’-methylenebisacrylamide (MBAA, cat. no. M7279), chlorophyllin sodium copper sodium salt (SCC, cat. no. C6003), 4,4’-azobis(4-cyanovaleric acid) (ACVA, cat. no.11590), thiazolyl blue tetrazolium bromide (MTT, cat. no. M5655), 5-fluorouracil (5-Fu, cat. no. F6627), fetal bovine serum (FBS, cat. no. T8153), 2-mercaptoehanol (cat. no. M3148) and trypan blue (cat. no. T8154) were purchased from Sigma-Aldrich (St. Louis, MO, USA). Sodium dodecyl sulfate (SDS, cat. no. 230421000) was purchased from Acros organics (Bridgewater, NJ, USA). gentamicin (cat. no. 15710-064) was purchased from Thermo Fisher Scientific (cat. no. F6627; Waltham, MA, USA). L929 cells were obtained from the Food Industry Research and Development Institute (Hsinchu, Taiwan) and maintained in α-MEM (cat. no. 11900-024, Gibco, Waltham, MA, USA) supplemented with 0.5% gentamicin, 0.04% 2-mercaptoehanol and 10% FBS.

Chang R, & Tsai W.B. (2018). Fabrication of Photothermo-Responsive Drug-Loaded Nanogel for Synergetic Cancer Therapy. Polymers, 10(10), 1098.

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2 mercaptoehanol

Lab products found in correlation

3 protocols using 2 mercaptoehanol

PDAC Organoids Potentiate CD8+ T Cell Killing

PDAC Organoids Potentiate CD8+ T Cell Killing

Synthesis and Characterization of Hydrogel Polymers

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