Revision 1

#52183Store at -20C

Cell Signaling Technology

Orders: 877-616-CELL (2355) [email protected]

Support: 877-678-TECH (8324)

Web: [email protected] cellsignal.com

3 Trask LaneDanversMassachusetts01923USA
For Research Use Only. Not for Use in Diagnostic Procedures.
Applications:

WB, IF-IC

REACTIVITY:

H M R

SENSITIVITY:

Endogenous

MW (kDa):

78

Source/Isotype:

Rabbit IgG

UniProt ID:

#P48506

Entrez-Gene Id:

2729

Product Information

Product Usage Information

Application Dilution
Western Blotting 1:1000
Immunofluorescence (Immunocytochemistry) 1:200 - 1:800

Storage

Supplied in 10 mM sodium HEPES (pH 7.5), 150 mM NaCl, 100 µg/mL BSA, 50% glycerol, and less than 0.02% sodium azide. Store at –20°C. Do not aliquot the antibody.

Specificity / Sensitivity

GCLC (E2Y7D) Rabbit mAb recognizes endogenous levels of total GCLC protein.

Species Reactivity:

Human, Mouse, Rat

Source / Purification

Monoclonal antibody is produced by immunizing animals with a synthetic peptide corresponding to residues surrounding Leu92 of human GCLC protein.

Background

Glutamate-cysteine ligase catalytic subunit (GCLC) catalyzes the ligation of glutamate and cysteine, the first and rate-limiting step in glutathione biosynthesis (1). Studies show that deubiquitinases protect cancer cells when glutathione synthesis is suppressed. Upon inhibition of both GCLC and deubiquitinases, cancer cells undergo proteotoxic stress, leading to cell death (2). In addition, deletion of Gclc enhances reactive oxygen species in mouse regulatory T cells (Tregs). Glutathione synthesized in part by Gclc is required for the suppressive function of mouse Tregs (3). Furthermore, in non-small cell lung cancer (NSCLC) cell lines, the transcription factor NRF2 stimulates GCLC expression. When NSCLC cells are starved for cysteine, GCLC, through its non-canonical activity, catalyzes the synthesis of γ-glutamyl-peptide, therefore reducing glutamate levels to protect cells against ferroptosis (1).

  1. Kang, Y.P. et al. (2021) Cell Metab 33, 174-189.e7.
  2. Harris, I.S. et al. (2019) Cell Metab 29, 1166-1181.e6.
  3. Kurniawan, H. et al. (2020) Cell Metab 31, 920-936.e7.

Species Reactivity

Species reactivity is determined by testing in at least one approved application (e.g., western blot).

Western Blot Buffer

IMPORTANT: For western blots, incubate membrane with diluted primary antibody in 5% w/v nonfat dry milk, 1X TBS, 0.1% Tween® 20 at 4°C with gentle shaking, overnight.

Applications Key

WB: Western Blotting IF-IC: Immunofluorescence (Immunocytochemistry)

Cross-Reactivity Key

H: human M: mouse R: rat Hm: hamster Mk: monkey Vir: virus Mi: mink C: chicken Dm: D. melanogaster X: Xenopus Z: zebrafish B: bovine Dg: dog Pg: pig Sc: S. cerevisiae Ce: C. elegans Hr: horse GP: Guinea Pig Rab: rabbit All: all species expected

Trademarks and Patents

Cell Signaling Technology is a trademark of Cell Signaling Technology, Inc.
All other trademarks are the property of their respective owners. Visit cellsignal.com/trademarks for more information.

限制使用

除非 CST 的合法授书代表以书面形式书行明确同意,否书以下条款适用于 CST、其关书方或分书商提供的书品。 任何书充本条款或与本条款不同的客书条款和条件,除非书 CST 的合法授书代表以书面形式书独接受, 否书均被拒书,并且无效。

专品专有“专供研究使用”的专专或专似的专专声明, 且未专得美国食品和专品管理局或其他外国或国内专管机专专专任何用途的批准、准专或专可。客专不得将任何专品用于任何专断或治专目的, 或以任何不符合专专声明的方式使用专品。CST 专售或专可的专品提供专作专最专用专的客专,且专用于研专用途。将专品用于专断、专防或治专目的, 或专专售(专独或作专专成)或其他商专目的而专专专品,均需要 CST 的专独专可。客专:(a) 不得专独或与其他材料专合向任何第三方出售、专可、 出借、捐专或以其他方式专专或提供任何专品,或使用专品制造任何商专专品,(b) 不得复制、修改、逆向工程、反专专、 反专专专品或以其他方式专专专专专品的基专专专或技专,或使用专品开专任何与 CST 的专品或服专专争的专品或服专, (c) 不得更改或专除专品上的任何商专、商品名称、徽专、专利或版专声明或专专,(d) 只能根据 CST 的专品专售条款和任何适用文档使用专品, (e) 专遵守客专与专品一起使用的任何第三方专品或服专的任何专可、服专条款或专似专专

Revision 1
#52183

GCLC (E2Y7D) Rabbit mAb

Western Blotting Image 1: GCLC (E2Y7D) Rabbit mAb Expand Image
使用 GCLC (E2Y7D) Rabbit mAb(上图)或 GAPDH (D16H11) XP® Rabbit mAb #5174(下图)对各种细胞系的提取物进行蛋白质印迹法分析。GCLC 蛋白在 A-172 细胞中低表达与预测的表达模式一致。
Western Blotting Image 2: GCLC (E2Y7D) Rabbit mAb Expand Image
使用 GCLC (E2Y7D) Rabbit mAb 对不同组织和细胞系的提取物进行蛋白质印迹法分析。
Western Blotting Image 3: GCLC (E2Y7D) Rabbit mAb Expand Image
使用 GCLC (E2Y7D) Rabbit mAb(上图)、DYKDDDDK Tag (9A3) Mouse mAb #8146(中图)或 GAPDH (D16H11) XP® Rabbit mAb #5174(下图),对转染空载 (-) 或转染带有全长 Myc/DDK 标签的人 GCLC 蛋白 (hGCLC-Myc/DDK; +) 的 293T 细胞提取物进行蛋白质印迹法分析。
Immunofluorescence Image 1: GCLC (E2Y7D) Rabbit mAb Expand Image
使用 GCLC (E2Y7D) Rabbit mAb(绿色)、DyLight 650 Phalloidin #12956(红色)和 DAPI #4083(蓝色)对 KYSE-70 细胞(左图,高表达)和 A-172 细胞(右图,低表达)进行共聚焦免疫荧光分析。