Caspase-3 is a widely expressed dimeric peptidase that is the major executioner caspase, the primary downstream mediator of apoptotic-associated proteolysis. Caspase-3 cleaves multiple substrates including PARP, proIL-16, PKC-gamma and -delta, proCaspases-6, -7, and -9, and beta-Catenin. Normally, it is an inactive, exclusively cytosolic homodimer. During apoptosis, however, proCaspase-3 is activated by cleavage into p20 and p12 subunits, and the p20 subunit is trimmed to yield a p17 subunit. Active Caspase-3 contains two p17 and two p12 subunits.
Human Active Caspase-3 Quantikine ELISA Kit
R&D Systems | Catalog # KM300
This product is in process of discontinuation.
Discontinued Product
KM300 has been discontinued.
View all Caspase-3 products.
Key Product Details
Assay Length
3.5 hours
Sample Type & Volume Required Per Well
Cell Extracts (100 µL)
Sensitivity
0.1 ng/mL
Assay Range
0.31-20 ng/mL (Cell Extracts)
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Product Summary for Human Active Caspase-3 Quantikine ELISA Kit
The Quantikine Human active Caspase-3 Immunoassay is a 3.5 hour solid phase ELISA designed to measure human active caspase-3 in cell extracts. It contains biotinylated-inhibitor derivatized recombinant caspase-3 and a monoclonal antibody raised against caspase-3. Results obtained using natural active caspase-3 covalently bound to biotin-ZVKD-fmk inhibitor in apoptotic cell extracts showed linear curves that were parallel to the Quantikine kit standard. These results indicate that this kit can be used to determine relative mass values for natural active caspase-3.
Product Specifications
Assay Type
Solid Phase Sandwich ELISA
Format
96-well strip plate
Measurement
Quantitative ELISA
Detection Method
Colorimetric - 450nm (TMB)
Conjugate
HRP
Species
Human
Specificity
Biotin-ZVKD-fmk modified natural and biotin-D-fmk modified recombinant active Caspase-3. This assay also detects mouse active Caspase-3.
Cross-reactivity
< 0.5% cross-reactivity observed with available related molecules. Cross-species reactivity observed with 1 or more species tested.
Interference
No significant interference observed with available related molecules.
Precision
Intra-Assay Precision (Precision within an assay) Three samples of known concentration were tested on one plate to assess intra-assay precision.
Inter-Assay Precision (Precision between assays) Three samples of known concentration were tested in separate assays to assess inter-assay precision.
Cell Extracts
| Intra-Assay Precision | Inter-Assay Precision | |||||
|---|---|---|---|---|---|---|
| Sample | 1 | 2 | 3 | 1 | 2 | 3 |
| n | 20 | 20 | 20 | 20 | 20 | 20 |
| Mean (pg/mL) | 1.06 | 3.09 | 7.70 | 1.06 | 3.27 | 8.25 |
| Standard Deviation | 0.05 | 0.16 | 0.20 | 0.07 | 0.19 | 0.50 |
| CV% | 4.7 | 5.2 | 2.6 | 6.6 | 5.8 | 6.1 |
Recovery for Human Active Caspase-3 Quantikine ELISA Kit
The recovery of active caspase-3 spiked to three levels throughout the range of the assay in various matrices was evaluated.
| Sample Type | Average % Recovery | Range % |
|---|---|---|
| Calibrator Diluent RD5-20 (1X) (n=4) | 99 | 98-100 |
| Extraction Buffer (1X) (n=4) | 110 | 104-114 |
| Extracts from Jurkat Cells Treated w/Staurosporine (n=4) | 92 | 87-95 |
Linearity
To assess the linearity of the assay, five or more samples containing and/or spiked with various concentrations of active caspase-3 in each matrix were diluted with Calibrator Diluent and then assayed. Results from typical sample dilutions are
Scientific Data Images for Human Active Caspase-3 Quantikine ELISA Kit
Human Active Caspase-3 ELISA Standard Curve
Preparation and Storage
Shipping
The product is shipped at ambient temperature. Upon receipt, store it immediately at the temperature recommended below.
Stability & Storage
Store the unopened product at 2 - 8 °C. Do not use past expiration date.
Background: Caspase-3
Alternate Names
Apopain, CASP3, Caspase3, CPP32, LICE-1, YAMA
Gene Symbol
CASP3
Additional Caspase-3 Products
Product Documents for Human Active Caspase-3 Quantikine ELISA Kit
Certificate of Analysis
To download a Certificate of Analysis, please enter a lot or batch number in the search box below.
Note: Certificate of Analysis not available for kit components.
Product Specific Notices for Human Active Caspase-3 Quantikine ELISA Kit
For research use only
⚠ WARNING: This product can expose you to chemicals including N,N-Dimethylforamide, which is known to the State of California to cause cancer. For more information, go to www.P65Warnings.ca.gov.Related Research Areas
Citations for Human Active Caspase-3 Quantikine ELISA Kit
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Protocols
View specific protocols for Human Active Caspase-3 Quantikine ELISA Kit (KM300):
Refer to the product for complete assay procedure.
Bring all reagents and samples to room temperature before use. It is recommended that all samples, standards, and controls be assayed in duplicate.
- Prepare all reagents, standard dilutions, and samples as directed in the product insert.
- Remove excess microplate strips from the plate frame, return them to the foil pouch containing the desiccant pack, and reseal.
- Add 100 µL of Standard, control, or sample to each well. Cover with a plate sealer, and incubate at room temperature for 2 hours.
- Aspirate each well and wash, repeating the process 4 times for a total of 5 washes.
- Add 100 µL of Conjugate to each well. Cover with a new plate sealer, and incubate at room temperature for 1 hour.
- Aspirate and wash 5 times.
- Add 100 µL Substrate Solution to each well. Incubate at room temperature for 30 minutes. PROTECT FROM LIGHT.
- Add 100 µL of Stop Solution to each well. Read at 450 nm within 30 minutes. Set wavelength correction to 540 nm or 570 nm.
100 µL Standard, Control, or Sample
100 µL Conjugate
100 µL Substrate Solution
100 µL Stop Solution
Find general support by application which include: protocols, troubleshooting, illustrated assays, videos and webinars.
- ELISA Sample Preparation & Collection Guide
- ELISA Troubleshooting Guide
- How to Run an R&D Systems DuoSet ELISA
- How to Run an R&D Systems Quantikine ELISA
- How to Run an R&D Systems Quantikine™ QuicKit™ ELISA
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- Quantikine HS ELISA Kit Principle, Streptavidin-HRP Polymer
- Sandwich ELISA (Colorimetric) – Biotin/Streptavidin Detection Protocol
- Sandwich ELISA (Colorimetric) – Direct Detection Protocol
- Troubleshooting Guide: ELISA
- View all Protocols, Troubleshooting, Illustrated assays and Webinars
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Associated Pathways
Apoptosis Signaling Pathway
