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59187
Beclin-1 Complex Antibody Sampler Kit
Primary Antibodies
Antibody Sampler Kit
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Beclin-1 Complex Antibody Sampler Kit #59187

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Beclin-1 Complex Antibody Sampler Kit: Image 1

Western blot analysis of extracts from various cell lines using Beclin-1 (D40C5) Rabbit mAb.

Beclin-1 Complex Antibody Sampler Kit: Image 2

Western blot analysis of extracts from mouse brain and C6 cells using PI3 Kinase Class III (D9A5) Rabbit mAb.

Beclin-1 Complex Antibody Sampler Kit: Image 3

Western blot analysis of extracts from various cell lines using PIK3R4 Antibody.

Beclin-1 Complex Antibody Sampler Kit: Image 4

Immunoprecipitation of Atg14 from HCT 116 cell extracts. Lane 1 is 10% input, lane 2 is precipitated with Rabbit (DA1E) mAb IgG XP® Isotype Control #3900, and lane 3 is Atg14 (D1A1N) Rabbit mAb. Western blot was performed using Atg14 (D1A1N) Rabbit mAb. A confirmation specific secondary antibody was used to avoid reactivity with IgG.

Beclin-1 Complex Antibody Sampler Kit: Image 5

Immunoprecipitation of UVRAG from PANC-1 cell extracts using Rabbit (DA1E) mAb IgG XP® Isotype Control #3900 (lane 2) or UVRAG (D2Q1Z) Rabbit mAb (lane 3). Lane 1 is 10% input. Western blot analysis was performed using UVRAG (D2Q1Z) Rabbit mAb.

Beclin-1 Complex Antibody Sampler Kit: Image 6

Western blot analysis of extracts from various cell lines using Rubicon (D9F7) Rabbit mAb.

Beclin-1 Complex Antibody Sampler Kit: Image 7

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.

Beclin-1 Complex Antibody Sampler Kit: Image 8

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.

Beclin-1 Complex Antibody Sampler Kit: Image 9

Western blot analysis of extracts from HCT 116 and HCT 116/Atg14 shRNA knockout cells using Atg14 (D1A1N) Rabbit mAb (upper) and β-Actin (D6A8) Rabbit mAb #84576 (lower). HCT 116/Atg14 shRNA cells were kindly provided by Dr. Do-Hyung Kim, University of Minnesota, Minneapolis, MN.

Beclin-1 Complex Antibody Sampler Kit: Image 10

Western blot analysis of extracts from various cell lines using UVRAG (D2Q1Z) Rabbit mAb (upper) or β-Actin (D6A8) Rabbit mAb #8457 (lower). HCT 116 cells have been reported to contain a monoallelic frameshift mutation resulting in significantly reduced levels of endogenous UVRAG expression (11).

Beclin-1 Complex Antibody Sampler Kit: Image 11

Western blot analysis of extracts from 293T cells, mock transfected (-) or transfected with human Rubicon (hRubicon, +), using Rubicon (D9F7) Rabbit mAb. Rubicon construct was kindly provided by Dr. Qing Zhong, University of California, Berkeley, CA.

Beclin-1 Complex Antibody Sampler Kit: Image 12

Western blot analysis of extracts from various cell lines using Atg14 (D1A1N) Rabbit mAb.

Beclin-1 Complex Antibody Sampler Kit: Image 13

Western blot analysis of extracts from 293T cells, mock transfected (-) or transfected with a construct expressing human UVRAG (hUVRAG; +), using UVRAG (D2Q1Z) Rabbit mAb.

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To Purchase # 59187T
Product # Size Price Inventory
59187T		 1 Kit  (6 x 20 µl)

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Product Description

The Beclin-1 Complex Antibody Sampler Kit provides an economical means of detecting proteins that are part of the Beclin-1 complexes. The kit includes enough antibodies to perform two western blot experiments with each primary antibody.

Specificity / Sensitivity

Each antibody in the Beclin-1 Complex Antibody Sampler Kit detects endogenous levels of its target protein. Rubicon (D9F7) Rabbit mAb detects a band of unknown origin at 55 kDa.

Source / Purification

Monoclonal antibodies are produced by immunizing animals with synthetic peptides corresponding to residues surrounding Thr72 of human Beclin-1, Lys630 of human PI3 Kinase Class III, Arg70 of human Atg14, Gly502 of human UVRAG, and Leu210 of human Rubicon. Polyclonal antibodies are produced by immunizing animals with a synthetic peptide corresponding to residues surrounding Gly825 of human PIK3R4 protein. Antibodies are purified by protein A and peptide affinity chromatography.

Background

A number of studies have identified distinct complexes involving Beclin-1 and PI3K Kinase Class III with specific roles in autophagy and vesicle trafficking (1,2). These complexes commonly contain Beclin-1, PI3KC3/VSP34, and PIK3R4/VPS15 and function to catalyze the phosphorylation of phosphatidylinositol at the D3 position, producing phosphatidylinositol-3-phosphate. Specificity of PI3KC3 activity is regulated by additional binding partners. Complex 1 contains Atg14 which is required for early stages of autophagosome nucleation (3,4). Complex 2 lacks Atg14, but instead contains UVRAG, and is important for autophagosome maturation and endocytic trafficking (4-6). A third complex, containing both UVRAG and Rubicon, negatively regulates canonical autophagy (7,8). Importantly, this complex containing Rubicon is critical for a related process of LC3-associated phagocytosis (LAP) in which extracellular pathogens binding to cell surface receptors are engulfed by a single membrane phagosome and degraded by the lysosome (9,10).

  1. Levine, B. and Kroemer, G. (2019) Cell 176, 11-42.
  2. Mei, Y. et al. (2016) Protein Sci 25, 1767-85.
  3. Sun, Q. et al. (2008) Proc Natl Acad Sci U S A 105, 19211-6.
  4. Itakura, E. et al. (2008) Mol Biol Cell 19, 5360-72.
  5. Liang, C. et al. (2006) Nat Cell Biol 8, 688-99.
  6. Liang, C. et al. (2008) Nat Cell Biol 10, 776-87.
  7. Matsunaga, K. et al. (2009) Nat Cell Biol 11, 385-96.
  8. Zhong, Y. et al. (2009) Nat Cell Biol 11, 468-76.
  9. Martinez, J. et al. (2015) Nat Cell Biol 17, 893-906.
  10. Heckmann, B.L. and Green, D.R. (2019) J Cell Sci 132(5). pii: jcs222984. doi: 10.1242/jcs.222984.

Pathways & Proteins

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