Product Pathways - Apoptosis / Autophagy

Cleaved Caspase-3 (Asp175) (5A1E) Rabbit mAb #9664

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Applications	Reactivity	Sensitivity	MW (kDa)	Isotype
W IP IHC-P IHC-F IF-IC F	H M R Mk	Endogenous	17, 19	Rabbit IgG

Applications Key: W=Western Blotting IP=Immunoprecipitation IHC-P=Immunohistochemistry (Paraffin) IHC-F=Immunohistochemistry (Frozen) IF-IC=Immunofluorescence (Immunocytochemistry) F=Flow Cytometry
Reactivity Key: H=Human M=Mouse R=Rat Mk=Monkey
Species cross-reactivity is determined by Western blot.

Protocols

9664:: Flow, IHC / Frozen, IHC / Paraffin, Immunofluorescence, Immunoprecipitation, Western Blotting

Specificity / Sensitivity

Cleaved Caspase-3 (Asp175) (5A1) Rabbit mAb detects endogenous levels of the large fragment (17/19 kDa) of activated caspase-3 resulting from cleavage adjacent to Asp175. This antibody does not recognize full length caspase-3 or other cleaved caspases.

Source / Purification

Monoclonal antibody is produced by immunizing animals with a synthetic peptide corresponding to amino-terminal residues adjacent to Asp175 of human caspase-3.

Western Blotting

Western blot analysis of extracts from C6 (rat), NIH/3T3 (mouse), and Jurkat (human) cells, untreated or treated with staurosporine (1uM, 3hrs) or etoposide (25uM, 5hrs) as indicated, using Cleaved Caspase-3 (Asp175) (5A1) Rabbit mAb.

IP

Immunoprecipitation of extracts from Jurkat cells, untreated or etoposide-treated (25uM, 5hrs), using Cleaved Caspase-3 (Asp175) (5A1) Rabbit mAb. Western blot was performed using the same antibody.

IHC-P (paraffin)

Immunofluorescent analysis of paraffin-embedded mouse embryo, using Cleaved Caspase-3 (Asp175) (5A1) Rabbit mAb (red) and DAPI (blue).

IHC-P (paraffin)

Immunohistochemical analysis of paraffin-embedded mouse embryo, using Cleaved Caspase-3 (Asp175) (5A1) Rabbit mAb in the presence of control peptide (left) or Cleaved Caspase-3 (Asp175) Blocking Peptide (#1050) (right).

IHC-P (paraffin)

Immunohistochemical analysis using Cleaved Caspase-3 (Asp175) (5A1) Rabbit mAb on SignalSlide (TM) Cleaved Caspase-3 IHC Controls #8104 (paraffin-embedded Jurkat cell, untreated (left) or etoposide-treated (right)).

IHC-P (paraffin)

Immunohistochemical staining of paraffin-embedded mouse embryo, showing cytoplasmic localization in apoptotic cells, using Cleaved Caspase-3 (Asp175) (5A1) Rabbit mAb.

IHC-F (frozen)

Immunohistochemical analysis of frozen H1650 xenograft, using Cleaved Caspase-3 (Asp175) (5A1) Rabbit mAb.

Flow Cytometry

Flow cytometric analysis of Jurkat cells, untreated (blue) or etoposide-treated (green), using Cleaved Caspase-3(Asp175) (5A1) Rabbit mAb compared to a nonspecific negative control antibody (red).

IF-IC

Confocal immunofluorescent images of HT-29 cells, untreated (left) or Staurosporine #9953 treated (right) labeled with Cleaved Caspase-3 (Asp175) (5A1) Rabbit mAb (green). Actin filaments have been labeled with Alexa Fluor® 555 phalloidin (red). Blue pseudocolor = DRAQ5™ (fluorescent DNA dye).

Background

Caspase-3 (CPP-32, Apoptain, Yama, SCA-1) is a critical executioner of apoptosis, as it is either partially or totally responsible for the proteolytic cleavage of many key proteins such as the nuclear enzyme poly (ADP-ribose) polymerase (PARP) (1). Activation of caspase-3 requires proteolytic processing of its inactive zymogen into activated p17 and p12 fragments. Cleavage of caspase-3 requires aspartic acid at the P1 position (2).

Application References

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Companion Products

Rabbit Monoclonals Produced Using Epitomics® Technology, U.S. Patent No. 5,675,063.

This product is intended for research purposes only. The product is not intended to be used for therapeutic or diagnostic purposes in humans or animals.