Description

The caspase family of cysteine proteases plays a key role in apoptosis and inflammation. ICE (interleukin-1ß converting enzyme) was the first member of this family to be discovered following a search for human proteins with homology to ced-3, a cell death gene identified in C. elegans. "Caspase" has been adopted as a root name for all family members based on the properties of these enzymes. The "c" reflects a cysteine protease mechanism and "aspase" refers to their ability to cleave after aspartic acid residues. Caspases are synthesized as inactive proenzymes that are processed in cells undergoing apoptosis by self-proteolysis and/or cleavage by another protease. The processed forms consist of large (17-22 kDa) and small (10-12 kDa) subunits which associate to form an active enzyme (34 kDa). Caspase-6 exists as two isoforms, designated Mch2α (caspase-6) and Mch2ß, a proteolytically inactive isoform which lacks half of the large subunit. Caspase-6 is most closely related to caspase-3, and can be proteolytically cleaved/activated by caspase-3 into large (p18) and small (p11) subunits. Similarly, caspase-3 has been shown to be activated by caspase-6, suggesting that these proteases may generate an amplification cycle during apoptosis. Other proteolytical targets are shown below in the Western blot analysis of caspase-6 (lane 1): Lamin A and a cytosolic nuclease which translocates to the Daudi B cell lysate after probing with anti-nucleus to promote DNA fragmentation. Caspase-6 has also been shown to cleave Fak (focal adhesion kinase), a non-receptor protein tyrosine kinase that transduces cell signals. Clone B93-4 identifies full length caspase-6 at survival and proliferation signals from contact sites between the cell surface and extracellular matrix. In this instance, caspase-6 cleavage inhibits the anti-apoptotic function of Fak. Therefore, the protease activity of caspase-6 plays both activating and inhibitory roles in apoptotic pathways. Caspase-6 is observed at ~34 kDa in SDS-PAGE. The antibody detects both full length caspase-6 (34 kDa), as well as the p11 subunit of the active enzyme. A synthetic peptide sequence corresponding to amino acids 271-285 (KKQVPCFASMLTKK) of human caspase-6 was used as immunogen.

Format

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Preparation and Storage

Store undiluted at 4°C.The monoclonal antibody was purified from tissue culture supernatant or ascites by affinity chromatography.

Product Notices

1. Since applications vary, each investigator should titrate the reagent to obtain optimal results.
2. Caution: Sodium azide yields highly toxic hydrazoic acid under acidic conditions. Dilute azide compounds in running water before discarding to avoid accumulation of potentially explosive deposits in plumbing.
3. Sodium azide is a reversible inhibitor of oxidative metabolism; therefore, antibody preparations containing this preservative agent must not be used in cell cultures nor injected into animals. Sodium azide may be removed by washing stained cells or plate-bound antibody or dialyzing soluble antibody in sodium azide-free buffer. Since endotoxin may also affect the results of functional studies, we recommend the NA/LE (No Azide/Low Endotoxin) antibody format, if available, for in vitro and in vivo use.
4. Please refer to www.bdbiosciences.com/pharmingen/protocols for technical protocols.