Background
The Bcl-2 family of apoptosis-related genes plays central roles in regulating apoptotic pathways (reviewed in Thomadaki and Scorilas, 2006). Regulation of cell death through apoptosis is critical for the maintenance of homeostasis, defense against infectious agents, and normal development. Bcl-2 family proteins regulate apoptosis primarily through the regulation of mitochondrial outer membrane permeability. In mammals, the family consists of both prosurvival (antiapoptotic) and proapoptotic (prodeath) members. Cellular homeostasis is thought to be dependent on a balance between the actions of prosurvival and proapoptotic proteins. Bcl-2 family proteins can be divided into 3 main subfamilies on the basis of their function and the content of their Bcl-2 homology (BH) domains, for example: 1) Prosurvival: Bcl-2, Bcl-XL, Bcl-W, A1, and Mcl-1 2) Proapoptotic (multidomain): Bax, Bak, and Bok. 3) BH3-only (proapoptotic): Bad, Bcl-XS, Bid, Bik, Bim, Blk, Bmf, Bnip, Noxa, and Puma. Prosurvival members inhibit cells from undergoing apoptosis, whereas proapoptotic and BH3-only subfamily members promote apoptosis. There are 4 BH domains (1-4) conserved among Bcl-2 family proteins. The BH domains are important for function as well as for heterodimerization between family members. Typical prosurvival family members have all four BH domains (1-4), whereas proapoptotic (multidomain) members have BH1, 2 and 3 domains and BH3-only members have only the BH3 domain. Overall, the relative ratio of prosurvival and proapoptotic proteins determines the suseptibility of a cell to various apoptotic stimuli. Many Bcl-2 family proteins are differentially expressed in various malignancies and some are useful prognostic biomarkers. Prosurvival proteins are often elevated in diverse cancers and have the potential to confer resistance to both endogenous cell death stimuli and cancer treatments. Alterations in the ratio or levels of Bcl-2 family proteins have been also associated with nonmalignant diseases including neurodegenerative diseases, autoimmune diseases, AIDs, Down’s syndrome, cardiovascular diseases, diabetes, glomerulonephritis, and muscular dystrophy. IMG-5695 recognizes Bid (~19-23 kDa) Bid.  It also recognizes cleaved/truncated forms which contain amino acids 28-55. The C-terminal cleaved/truncated form (11-15 kDa), often referred as tBid in the literature, is not predicted to contain amino acids 28-55 (reviewed in Yin, 2006).     Howwever, IMG-5694 may recognize tBid in cases where  the smaller N-terminal portion (containing amino acids 1-55) of Bid has not separated from the larger C-terminal portion.  Please see Applications Notes for more information on the biology of Bid.  

Antigen
A synthetic peptide corresponding to amino acids 28-50 (CSDNSFRRELDALGHELPVLAPQ) of human Bid (also known as Bid, isoform 2) was used as immunogen; GenBank no. NP_001187.1. Human Bid is a 195 amino acid protein.

Application Notes
Full-length Bid is known to undergo cleavage/truncation (reviewed in Yin, 2006).  Bid was initially found to be cleaved and activated by capspase-8 following death receptor activation.  The term tBid was first used to described the caspase-8 cleaved/truncated C-terminal Bid.  Bid can be also cleaved by other proteases such as Granzyme B, calpains and cathepsin.  The C-terminal portion is considered to be the active Bid moiety.  This active form can translocate from the cytosol to the mitochondria. Therefore the appearance of Bid in the mitochondria is considered to be an indication of active Bid.  However, it should also be noted that Bid has been shown to translocate to the mitochondria without cleavage in some model systems.  The proteolytic cleavage of Bid usually occurs in the unstructured loop region between the alpha 2 and alpha 3 helices, which is between amino acids (aa) 41 and 79 of Bid. For example, the caspase-8/3 cleavage site is at 60 (human) and 59 (mouse).  It should be noted that after cleavage, the smaller N-terminal portion of Bid is not necessarily separated from the larger C-terminal portion.  The exact size of the C-terminal cleavage products depends on the Bid cleavage sites; generally 11-15 kDa C-truncated cleavage forms are generated.  Shorter C-terminal Bid cleavage products may result from protease cleavage that involves both caspases and other proteases. Full length Bid is quite stable, but cleaved/truncated forms may have a short half-life (less than 1.5 h) and degrade rapidly.  Therefore a time course may be useful when detecting Bid cleavage fragments. 

Genebank Info (Protein)
NP_001187

Related Products

Reference
1. Thomadaki H and A Scorilas. 2006. Bcl2 family of apoptosis-related genes: Functions and clinical implications in cancer. Critical Reviews in Clinical Laboratory Sciences. 43:1-67.
2. Yin. 2006. Bid, a BH3-only multi-functional molecule, is at the cross road of life and death. Gene. 369:7-19.

Product Citations
1.

Early processing of Bid and caspase-6, -8, -10, -14 in the canine brain during cardiac arrest and resucitation. Krajewska M, RE Rosenthal, J Mikolajczyk, HR Stennicke, T Wiesenthal, J Mai, M Naito, GS Salvesen, JC Reed, G Fiskum and S Krajewski. Experimental Neurology 189:261-279 (2004). 
Imgenex antibodies collectively cited for WB and IHC (P) on mouse spleen and thymus lysates, dog and rat brain tissue  and lysates, tissue culture transfectants, caspase-9 knockout mice, recombinant caspase proteins. 
Note:  Antibody applications and testing material vary, please see  the publication for additional details about how the antibodies were used.
1. Bak: IMG-5679
2. Bax: IMG-5683
3. Bcl-2: IMG-5685
4. Bcl-2: IMG-5686
5. Bcl-2: IMG-5687
6. Bcl-2: IMG-5688
7. Bcl-XL+S: IMG-5691
8. Bcl-XL+S: IMG-5692
9. Bid: IMG-5693
10. Bid: IMG-5694
11. Bid: IMG-5695
12. Caspase-3: IMG-5699
13. Caspase-6 (Active/Cleaved): IMG-5701
14. Caspase-7 (Active/Cleaved): IMG-5702
15. Caspase-8 (Active/Cleaved): IMG-5703 
16. Caspase-9 (Active/Cleaved): IMG-5705
17. Caspase-9: IMG-5706
18. Caspase-9: IMG-5707 
19. Caspase-10: IMG-5710
20. Caspase-10: IMG-5711
21. Caspase-10: IMG-5712
22. Caspase-14: IMG-5713