Apoptosis or programmed cell death is a fundamental cellular process that is essential for normal tissue development and abnormal growth.
Apoptosis is driven by two classes of specialized proteases known as caspases (Cysteine Aspartase).
The initiator caspases can be activated by self-cleavage.
The effector caspases are then activated in an amplification cascade.
Several key factors are released from the mitochondria that regulate apoptosis.
The first such factor (Cytochrome-C) to be described binds to a cytoplasmic scaffolding protein called Apaf-1 (Apoptosis Protease activating factor-1), a homolog of C.
elegans Ced-4.
Both Apaf-1 and Ced-4 are composed of an N-terminal Caspase Recrutiment domain (CARD) linked to a Nuceoltide-bidning domain (NBD), also known as NB-ARC or NOD domain.
Ced-4 and Apaf-1 self-associate via the NBD and activate Casp-3 and -9.
The C-terminal region of Apaf-1 lacks homology with Ced-4 and contains 12 WD-40 repeats.
IN response to certain apoptotic stimuli, Cytochrome-C is released from the mitochondria and binds to Apaf-1 to form a ternary complex with, and activate, the initiator pro-caspase-9.
Active caspase-9 then turns on downstream effector caspases, initiating apoptosis.
The presence of CARD motifs in a variety of other effector and signaling molecules, including numerous caspases, NF- B-activating kinase, suggests that other CED-4/Apaf-1 family members likely exist in humans to coordinate downstream stress responses.
Recently, Additional members of Apaf-1 family, NOD1/CARD4 and NOD2 have been cloned and characterized.
Apaf-1 (human 1205 aa; chromosome 12q23; mosue 1238 aa; rat 1249 aa; mol wt ~130kD) has significant (~20%) homology with ced-3 and ced-4.
It is a cytoplsamic protein found in most tissues with highest expression in adult spleen and peripheral blood leukocytes, fetal brain, kidney, and lung correlating with high level of apoptosis in these tissues.
Apaf-1 participates in the cytochrome-c dependent activation of caspase-3.
The N-terminal 85-aa of Apaf-1 show 21% identity with C.
elegans CED3.
The next 320-aa show ~22% idneitty with CED4.
C-terminus of Apaf-1 comprised of 12 WD repeats (TRP-Asp domains), which mediate proteinpprotein interactions.
Apaf-1 is alternatively spliced to three forms that vary in length.
Applications:Suitable for use in ELISA, Western Blotting.
Other applications not tested.
Recommended Dilution:Western Blotting ~1.0ug/ml for affinity pure antibody using ECL.
Apaf-1 is ~130kD
ELISA: 1:10,000-50,000 for neat serum and 0.5-1ug/ml for affinity pure.
Control peptide can be used to coat ELISA plates at 1ug/ml.
Optimal dilutions to be determined by the researcher.
Storage and Stability:May be stored at 4°C for short-term only.
For long-term storage and to avoid repeated freezing and thawing, aliquot and add glycerol (40-50%).
Freeze at -20°C.
Aliquots are stable for at least 12 months at -20°C.
For maximum recovery of product, centrifuge the original vial after thawing and prior to removing the cap.
Further dilutions can be made in assay buffer.
仕様
Size：100ug
Host：rabbit
Source Antibody：human
Grade：Affinity Purified
Purity：Purified by immunoaffinity chromatography.
Form：Supplied as a liquid in PBS, pH 7.4 and 0.09% sodium azide, 40% glycerol.
Specificity：Human is 100% conserved in mouse and 94% in rat Apaf-1. It has no significant homology with ced-3/4, Nod-1 and NOD-2 family of proteins. Control peptide, because of its low MW (<3kD), is not suitable for Western. It should be used for ELISA or antibody blocking experiments (use 5-10 ug control peptide per 1 ug of aff pure IgG or 1 ul antiserum) to confirm antibody specificity
