Revision 2

#4445

Store at -20C

Autophagy Antibody Sampler Kit

1 Kit

(7 x 20 microliters)

CST Logo
Orders:

877-616-CELL (2355)

[email protected]

Support:

877-678-TECH (8324)

3 Trask Lane | Danvers | Massachusetts | 01923 | USA

For Research Use Only. Not for Use in Diagnostic Procedures.

Product Includes Product # Quantity Mol. Wt Isotype/Source
Beclin-1 (D40C5) Rabbit mAb349520 µl60 kDaRabbit IgG
LC3A/B (D3U4C) XP® Rabbit mAb1274120 µl14, 16 kDaRabbit IgG
Atg5 (D5F5U) Rabbit mAb1299420 µl55 kDaRabbit IgG
Atg12 (D88H11) Rabbit mAb418020 µl16, 55 kDaRabbit IgG
Atg16L1 (D6D5) Rabbit mAb808920 µl66, 68 kDaRabbit IgG
Atg7 (D12B11) Rabbit mAb855820 µl78 kDaRabbit IgG
Atg3 Antibody341520 µl40 kDaRabbit
Anti-rabbit IgG, HRP-linked Antibody7074100 µlGoat

Please visit cellsignal.com for individual component applications, species cross-reactivity, dilutions, protocols, and additional product information.

Description

The Autophagy Antibody Sampler Kit provides an economical means to investigate the molecular machinery of autophagy within the cell. The kit contains enough primary and secondary antibodies to perform two Western blot experiments.

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.

Background

Autophagy is a catabolic process for the autophagosomic-lysosomal degradation of bulk cytoplasmic contents (1,2). Autophagy is generally activated by conditions of nutrient deprivation but has also been associated with a number of physiological processes including development, differentiation, neurodegeneration, infection, and cancer (3). The molecular machinery of autophagy was largely discovered in yeast and referred to as autophagy-related (Atg) genes. Formation of the autophagosome involves a ubiquitin-like conjugation system in which Atg12 is covalently bound to Atg5 and targeted to autophagosome vesicles (4-6). This conjugation reaction is mediated by the ubiquitin E1-like enzyme Atg7 and the E2-like enzyme Atg10 (7,8).

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

Orders: 877-616-CELL (2355) [email protected] Support: 877-678-TECH (8324) [email protected] Web: cellsignal.com
For Research Use Only. Not for Use in Diagnostic Procedures.

Revision 2

Cell Signaling Technology Logo
Western blot analysis of extracts from RD cells, untreated (-) or Torin 1-treated (250 nM, 4 hr; +), using LC3A/B (D3U4C) XP® Rabbit mAb.
undefined Image 1: Autophagy Antibody Sampler Kit
Western blot analysis of extracts from various cell lines using Atg5 (D5F5U) Rabbit mAb.
undefined Image 2: Autophagy Antibody Sampler Kit
Western blot analysis of extracts from various cell lines using Atg3 Antibody.
undefined Image 3: Autophagy Antibody Sampler Kit
Orders: 877-616-CELL (2355) [email protected] Support: 877-678-TECH (8324) [email protected] Web: cellsignal.com
For Research Use Only. Not for Use in Diagnostic Procedures.

Revision 2

Cell Signaling Technology Logo
Western blot analysis of extracts from various cell lines using Beclin-1 (D40C5) Rabbit mAb.
undefined Image 4: Autophagy Antibody Sampler Kit
Western blot analysis of extracts from various cell lines using Atg12 (D88H11) Rabbit mAb.
undefined Image 5: Autophagy Antibody Sampler Kit
After the primary antibody is bound to the target protein, a complex with HRP-linked secondary antibody is formed. The LumiGLO® is added and emits light during enzyme catalyzed decomposition.
undefined Image 6: Autophagy Antibody Sampler Kit
Orders: 877-616-CELL (2355) [email protected] Support: 877-678-TECH (8324) [email protected] Web: cellsignal.com
For Research Use Only. Not for Use in Diagnostic Procedures.

Revision 2

Cell Signaling Technology Logo
Western blot analysis of extracts from various cell lines using Atg16L1 (D6D5) Rabbit mAb.
undefined Image 7: Autophagy Antibody Sampler Kit
Western blot analysis of extracts from various cell lines using Atg7 (D12B11) Rabbit mAb.
undefined Image 8: Autophagy Antibody Sampler Kit
Western blot analysis of extracts from HeLa, NIH/3T3, and KNRK cells, untreated (-) or chloroquine-treated (50 μM, overnight; +), using LC3A/B (D3U4C) XP® Rabbit mAb.
undefined Image 9: Autophagy Antibody Sampler Kit
Orders: 877-616-CELL (2355) [email protected] Support: 877-678-TECH (8324) [email protected] Web: cellsignal.com
For Research Use Only. Not for Use in Diagnostic Procedures.

Revision 2

Cell Signaling Technology Logo
Western blot analysis of extracts from wild-type MEFs (wt) or MEFs from Atg5 knockouts (Atg5-/-) using Atg5 (D5F5U) Rabbit mAb (upper), or β-Actin (D6A8) Rabbit mAb #8457 (lower). Atg5-/- MEFs were kindly provided by Dr. Ramnik Xavier, Massachusetts General Hospital, Harvard Medical School, Boston, MA.
undefined Image 10: Autophagy Antibody Sampler Kit
Immunoprecipitation of Atg5 from PANC-1 cell extracts using Rabbit (DA1E) mAb IgG XP® Isotype Control #3900 (lane 2) or Atg5 (D5F5U) Rabbit mAb (lane 3). Lane 1 is 10% input. Western blot was performed using Atg5 (D5F5U) Rabbit mAb. Mouse Anti-rabbit IgG (Conformation Specific) (L27A9) mAb #3678 was used as a secondary antibody to avoid cross-reactivity with rabbit IgG.
undefined Image 11: Autophagy Antibody Sampler Kit
Immunohistochemical analysis of paraffin-embedded human squamous cell lung carcinoma using LC3A/B (D3U4C) XP® Rabbit mAb in the presence of control peptide (left) or antigen-specific peptide (right).
undefined Image 12: Autophagy Antibody Sampler Kit
Orders: 877-616-CELL (2355) [email protected] Support: 877-678-TECH (8324) [email protected] Web: cellsignal.com
For Research Use Only. Not for Use in Diagnostic Procedures.

Revision 2

Cell Signaling Technology Logo
Confocal immunofluorescent analysis of HeLa (upper) and C2C12 (lower) cells, chloroquine-treated (50 μM, overnight; left), nutrient-starved with EBSS (3 hr, middle) or untreated (right) using LC3A/B (D3U4C) XP® Rabbit mAb (green) and β-Actin (13E5) Rabbit mAb (Alexa Fluor® 555 Conjugate) #8046 (red). Blue pseudocolor= DRAQ5® #4084 (fluorescent DNA dye).
undefined Image 13: Autophagy Antibody Sampler Kit
Flow cytometric analysis of HeLa cells, untreated (blue) or treated with chloroquine (50 µM, 16 hr; green) using LC3A/B (D3U4C) Rabbit mAb. Anti-rabbit IgG (H+L), F(ab')2 Fragment (Alexa Fluor® 647 Conjugate) #4414 was used as a secondary antibody.
undefined Image 14: Autophagy Antibody Sampler Kit
Simple WesternTM analysis of lysates (0.1 mg/mL) from HeLa cells using Atg5 (D5F5U) Rabbit mAb #12994. The virtual lane view (left) shows the target band (as indicated) at 1:50 and 1:250 dilutions of primary antibody. The corresponding electropherogram view (right) plots chemiluminescence by molecular weight along the capillary at 1:50 (blue line) and 1:250 (green line) dilutions of primary antibody. This experiment was performed under reducing conditions on the JessTM Simple Western instrument from ProteinSimple, a BioTechne brand, using the 12-230 kDa separation module.
undefined Image 15: Autophagy Antibody Sampler Kit
Orders: 877-616-CELL (2355) [email protected] Support: 877-678-TECH (8324) [email protected] Web: cellsignal.com
For Research Use Only. Not for Use in Diagnostic Procedures.

Revision 2

Cell Signaling Technology Logo
Western blot analysis of extracts from HeLa cells, mock transfected or transfected with mouse Atg3, using Atg3 Antibody.
undefined Image 16: Autophagy Antibody Sampler Kit
Western blot analysis of extracts from HeLa cells, transfected with 100 nM SignalSilence® Control siRNA (Unconjugated) #6568 (-), SignalSilence® Beclin-1 siRNA I #6222 (+) or SignalSilence® Beclin-1 siRNA II (+), using Beclin-1 (D40C5) XP® Rabbit mAb #3495 (upper) or α-Tubulin (11H10) Rabbit mAb #2125 (lower). The Beclin-1 (D40C5) XP® Rabbit mAb confirms silencing of Beclin-1 expression, while the α-Tubulin (11H10) Rabbit mAb is used to control for loading and specificity of Beclin-1 siRNA.
undefined Image 17: Autophagy Antibody Sampler Kit
Western blot analysis of extracts from 293T cells, mock transfected (-) or transfected with either a myc-tagged human Atg16L1β construct (hAtg16L1β-Myc; +) or a mouse Atg16L1α construct (mAtg16L1α; +), using Atg16L1 (D6D5) Rabbit mAb. The myc-tagged human Atg16L1β construct was kindly provided by Dr. Qing Zhong, University of California Berkeley.
undefined Image 18: Autophagy Antibody Sampler Kit
Orders: 877-616-CELL (2355) [email protected] Support: 877-678-TECH (8324) [email protected] Web: cellsignal.com
For Research Use Only. Not for Use in Diagnostic Procedures.

Revision 2

Cell Signaling Technology Logo
Western blot analysis of extracts from HeLa cells, transfected with 100 nM SignalSilence® Control siRNA (Unconjugated) #6568 (-) or SignalSilence® Atg7 siRNA I #6604 (+), using Atg7 (D12B11) Rabbit mAb (upper) or α-Tubulin (11Η10) Rabbit mAb #2125 (lower). The Atg7 (D12B11) Rabbit mAb confirms silencing of Atg7 expression, while the α-Tubulin (11H10) Rabbit mAb is used as a loading control.
undefined Image 19: Autophagy Antibody Sampler Kit
Immunohistochemical analysis of paraffin-embedded mouse prostate using LC3A/B (D3U4C) XP® Rabbit mAb.
undefined Image 20: Autophagy Antibody Sampler Kit
Confocal immunofluorescent analysis of EBSS-starved PANC-1 cells using Atg16L1 (D6D5) Rabbit mAb (green). Blue pseudocolor = DRAQ5® #4084 (fluorescent DNA dye).
undefined Image 21: Autophagy Antibody Sampler Kit
Orders: 877-616-CELL (2355) [email protected] Support: 877-678-TECH (8324) [email protected] Web: cellsignal.com
For Research Use Only. Not for Use in Diagnostic Procedures.

Revision 2

Cell Signaling Technology Logo
Immunohistochemical analysis of paraffin-embedded NIH/3T3 cell pellets, control (left) or chloroquine-treated (right), using LC3A/B (D3U4C) XP® Rabbit mAb.
undefined Image 22: Autophagy Antibody Sampler Kit
Immunoprecipitation of Atg16L1 from Jurkat cell extracts. Lane 1 is 10% input, lane 2 is precipitated with Rabbit (DA1E) mAb IgG XP® Isotype Control #3900, and lane 3 is Atg16L1 (D6D5) Rabbit mAb, #8089. Western blot was performed using Atg16L1 (D6D5) Rabbit mAb.
undefined Image 23: Autophagy Antibody Sampler Kit
Orders: 877-616-CELL (2355) [email protected] Support: 877-678-TECH (8324) [email protected] Web: cellsignal.com
For Research Use Only. Not for Use in Diagnostic Procedures.