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| Product Name | Caspase-9 Activity Colorimetric Assay Kit |
| Catalog No. | CCAT-HMM-0056 |
| Description | A colorimetric assay kit for measuring caspase-9 activity, the key initiator caspase of the intrinsic mitochondrial apoptosis pathway. The assay employs an LEHD-p-nitroanilide chromogenic substrate that, upon cleavage by active caspase-9, releases p-nitroaniline detected at 405 nm. This kit enables quantitative comparison of caspase-9 activation across different treatment conditions. |
| Intended Use | Quantification of caspase-9 protease activity in cell lysates as a readout of intrinsic mitochondrial apoptotic pathway activation. |
| Principle / Technology | Chromogenic LEHD-pNA substrate cleavage by active caspase-9 |
| Detection Method | Colorimetric (absorbance at 405 nm) |
| Sample Type | Cell lysates and tissue homogenates from apoptotic cells induced via mitochondrial pathway |
| Performance Range / Specifications | Linear range: 0.02–2.0 absorbance units after 2 hr incubation at 37 °C |
| Sensitivity / LOD | Detection limit: 0.01 absorbance units above blank |
| Specificity | LEHD recognition sequence preferentially cleaved by caspase-9; inhibitor LEHD-fmk blocks ≥90% of activity |
| Reaction Conditions / Protocol | Induce apoptosis; lyse cells 10 min on ice with provided buffer; incubate lysate with LEHD-pNA substrate 1–2 hr at 37 °C; read at 405 nm |
| Components / Formulation | LEHD-pNA chromogenic substrate (lyophilized), pNA standard, caspase-9 reaction buffer, cell lysis buffer, DTT solution, LEHD-fmk inhibitor |
| Storage Conditions | –20 °C for substrate and inhibitor; 2–8 °C for buffers |
| Shelf Life | 12 months at –20 °C |
| Package Specifications | 100 tests |
| Product Form | Lyophilized substrate; liquid buffers |
| Quality Control | Each lot validated with cytochrome c/dATP-activated cell lysate; inhibitor reduces signal to baseline |
| Key Features | Colorimetric format enables detection on standard microplate readers without fluorescence capability |
| Purity | Reagent grade; fluorophore-labeled reagents ≥90% labeling efficiency |
| Concentration | As specified per kit; sufficient for stated number of tests |
| Activity / Unit Definition | Apoptosis induction confirmed by positive control treatment producing ≥5-fold signal increase |
| Molecular Weight | Varies by dye and enzyme component |
| Source / Origin | Recombinant annexin proteins; synthetic fluorophore conjugates; chromogenic/fluorogenic peptide substrates |
| pH Range / Optimal pH | Binding/washing buffer pH 7.2–7.4 |
| Shipping Conditions | Cold packs; protect fluorophores from light; some components may require –20 °C |
| Expiration Date / Stability | 6–12 months at recommended storage temperature |
| Regulatory / Compliance | Research use; manufactured under ISO 9001 |
| Compatibility | Compatible with flow cytometry and fluorescence microscopy platforms |
| Recommended Buffer System | Annexin binding buffer with calcium; cell lysis buffer with protease inhibitors for caspase assays |
| Application Notes / Precautions | Process samples promptly after staining; include unstained and single-stain controls for flow cytometry compensation; use appropriate apoptosis inducers as positive controls |
| Batch-to-Batch Consistency | Positive control signal within ±20% of reference lot; staining pattern consistent |
For research use only, not for clinical use.
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