Caspase9 Antibody [A3K12] | Primary Antibodies

Application: Reactivity: Human, Mouse, Rat, Hamster, Monkey

Lane 1: Jurkat, Lane 2: Jurkat (staurosporine treated)

Usage Information

Dilution
WB1:1000

Application
WB

Reactivity
Human, Mouse, Rat, Hamster, Monkey

Source
Mouse Monoclonal Antibody

Storage Buffer
PBS, pH 7.2+50% Glycerol+0.05% BSA+0.01% NaN3

Storage (from the date of receipt)
-20°C (avoid freeze-thaw cycles), 2 years

Predicted MW
35-51 kDa

Datasheet & SDS

Biological Description

Specificity
Caspase9 Antibody [A3K12] detects endogenous levels of total Caspase9 protein.

Clone
A3K12

Synonym(s)
Caspase-9; CASP-9; Apoptotic protease Mch-6; Apoptotic protease-activating factor 3 (APAF-3); ICE-like apoptotic protease 6 (ICE-LAP6); CASP9; MCH6

Background
Caspase-9 is an initiator caspase central to the intrinsic apoptosis pathway, acting at the apex of the mitochondrial death signaling cascade. Upon cytochrome c release from mitochondria, caspase-9 assembles into the Apaf-1 apoptosome. This process involves induced proximity oligomerization via Apaf-1 CARD domain interactions, leading to autocatalytic cleavage at Asp315 and formation of an active p35/p12 heterodimer. A secondary cleavage at Asp330 yields a p37 fragment, further amplifying effector caspase activation. The apoptosome-bound caspase-9 demonstrates over 100-fold enhanced activity due to allosteric stabilization of its active conformation, maintained by Apaf-1 even without continued dATP presence after assembly. This holoenzyme efficiently activates caspase-3 and caspase-7, initiating the executioner phase, which results in PARP cleavage, DNA fragmentation, and cytoskeletal collapse. Physiological inhibition occurs through Xiap, which binds the active site and BIR2 domain of processed caspase-9; however, caspase-3 feedback cleavage of Xiap relieves this inhibition, ensuring irreversible apoptosis commitment. Alternative splicing produces the dominant-negative caspase-9S isoform, which lacks catalytic activity and competes for apoptosome docking. Caspase-9 is essential for developmental sculpting, DNA damage surveillance, and stress-induced clearance in neurons and lymphocytes. It serves as the canonical marker for intrinsic apoptosis in research assays using fluorogenic substrates or dominant-negative mutants in flow cytometry. Caspase-9 deficiency leads to severe lymphoproliferation and autoimmunity, while APAF-1 methylation in cancer silences the pathway.

Background

Caspase-9 is an initiator caspase central to the intrinsic apoptosis pathway, acting at the apex of the mitochondrial death signaling cascade. Upon cytochrome c release from mitochondria, caspase-9 assembles into the Apaf-1 apoptosome. This process involves induced proximity oligomerization via Apaf-1 CARD domain interactions, leading to autocatalytic cleavage at Asp315 and formation of an active p35/p12 heterodimer. A secondary cleavage at Asp330 yields a p37 fragment, further amplifying effector caspase activation. The apoptosome-bound caspase-9 demonstrates over 100-fold enhanced activity due to allosteric stabilization of its active conformation, maintained by Apaf-1 even without continued dATP presence after assembly. This holoenzyme efficiently activates caspase-3 and caspase-7, initiating the executioner phase, which results in PARP cleavage, DNA fragmentation, and cytoskeletal collapse. Physiological inhibition occurs through Xiap, which binds the active site and BIR2 domain of processed caspase-9; however, caspase-3 feedback cleavage of Xiap relieves this inhibition, ensuring irreversible apoptosis commitment. Alternative splicing produces the dominant-negative caspase-9S isoform, which lacks catalytic activity and competes for apoptosome docking. Caspase-9 is essential for developmental sculpting, DNA damage surveillance, and stress-induced clearance in neurons and lymphocytes. It serves as the canonical marker for intrinsic apoptosis in research assays using fluorogenic substrates or dominant-negative mutants in flow cytometry. Caspase-9 deficiency leads to severe lymphoproliferation and autoimmunity, while APAF-1 methylation in cancer silences the pathway.

References
https://pubmed.ncbi.nlm.nih.gov/10617566/ https://pubmed.ncbi.nlm.nih.gov/11904389/

Reference Table for Selecting PVDF Membrane Pore Size Specifications

Protein Size (kDa)	PVDF Transfer Membrane Pore Size (μm)
< 10	0.22
10-20	0.22
20-30	0.45
30-45	0.45
45-70	0.45
80-100	0.45
100-140	0.45
> 140	0.45

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Protein Size (kDa)	Separating Gel Percentage
4-40	20%
12-45	15%
10-70	12.5%
15-100	10%
25-100	8%
60-210	5%