Product Pathways - Autophagy Signaling
Autophagy Antibody Sampler Kit #4445
|Kit Includes||Quantity||Applications||Reactivity||MW (kDa)||Isotype|
|Beclin-1 (D40C5) Rabbit mAb #3495||40 µl||W IP||H M R Mk||60||Rabbit IgG|
|LC3A (D50G8) XP® Rabbit mAb #4599||40 µl||W IHC-P IF-IC F||H M R (Mk) (Dg)||14, 16||Rabbit IgG|
|LC3B (D11) XP® Rabbit mAb #3868||40 µl||W IP IHC-P IF-IC F||H M R (Mk) (B) (Pg)||14, 16||Rabbit IgG|
|Atg5 (D1G9) Rabbit mAb #8540||40 µl||W IP||H M R Mk (B) (Dg) (Pg)||55||Rabbit IgG|
|Atg12 (D88H11) Rabbit mAb #4180||40 µl||W IP||H M R Mk||16, 55||Rabbit IgG|
|Atg7 (D12B11) Rabbit mAb #8558||40 µl||W IP||H M R (Mk) (B)||78||Rabbit IgG|
|Atg3 Antibody #3415||40 µl||W||H M R Mk (C) (X) (B) (Dg)||40||Rabbit|
|Anti-rabbit IgG, HRP-linked Antibody #7074||100 µl||Goat|
Reactivity Key: H=Human M=Mouse R=Rat Mk=Monkey C=Chicken X=Xenopus B=Bovine Dg=Dog Pg=Pig
Species enclosed in parentheses are predicted to react based on 100% sequence homology.
Specificity / Sensitivity
Each antibody in the Autophagy Antibody Sampler Kit detects its respective target at endogenous levels. Both the Atg5 and Atg12 rabbit monoclonal antibodies recognize the Atg12-Atg5 conjugate. The LC3A and LC3B rabbit monoclonal antibodies may cross-react with other LC3 isoforms. The LC3B rabbit monoclonal antibody has stronger reactivity with the type II form of LC3B by western immunoblot.
Western blot analysis of extracts from various cell lines using Atg3 Antibody #3415.
Western blot analysis of extracts from various cell lines using Beclin-1 (D40C5) Rabbit mAb #3495.
Western blot analysis of extracts from HeLa cells, untreated or chloroquine-treated (50 μM, overnight), using LC3B (D11) XP® Rabbit mAb #3868.
Western blot analysis of extracts from various cell lines using Atg12 (D88H11) Rabbit mAb #4180.
Western blot analysis of extracts from HeLa and NIH/3T3 cells, untreated or chloroquine-treated (50 μM, overnight), using LC3A (D50G8) XP® Rabbit mAb #4599.
Immunoprecipitation of Atg5 from HT-1080 cells using Atg5 (D1G9) Rabbit mAb #8540. Western blot detection was performed using the same antibody (upper), or with Atg12 (D88H11) Rabbit mAb #4180 (lower). Lane 1 is 10% input, lane 2 is precipitated with non-specific rabbit IgG, lane 3 is precipitated with Atg5 (D1G9) Rabbit mAb #8540.
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 four Western blot experiments.
Source / Purification
Monoclonal antibodies are produced by immunizing animals with synthetic peptides corresponding to residues surrounding Thr72 of human Beclin-1, Pro187 of human Atg5, Ser36 of human Atg12, or near the amino termini of human LC3A, LC3B, and Atg7. Polyclonal antibodies are produced by immunizing animals with synthetic peptides corresponding to residues near the amino-termini of human Atg3. Polyclonal antibodies are purified by protein A and peptide affinity chromatography.
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).
- Reggiori, F. and Klionsky, D.J. (2002) Eukaryot Cell 1, 11-21.
- Codogno, P. and Meijer, A.J. (2005) Cell Death Differ 12 Suppl 2, 1509-18.
- Levine, B. and Yuan, J. (2005) J Clin Invest 115, 2679-88.
- Mizushima, N. et al. (1998) J Biol Chem 273, 33889-92.
- Mizushima, N. et al. (1998) Nature 395, 395-8.
- Suzuki, K. et al. (2001) EMBO J 20, 5971-81.
- Tanida, I. et al. (1999) Mol Biol Cell 10, 1367-79.
- Shintani, T. et al. (1999) EMBO J 18, 5234-41.
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* Product-specific protocol.
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- 7074 Anti-rabbit IgG, HRP-linked Antibody
- 7720 Prestained Protein Marker, Broad Range (Premixed Format)
- 7727 Biotinylated Protein Ladder Detection Pack
- 7003 20X LumiGLO® Reagent and 20X Peroxide
For Research Use Only. Not For Use In Diagnostic Procedures.