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| Product Name | JC-1 Mitochondrial Membrane Potential Assay Kit |
| Catalog No. | CCAT-HMM-0047 |
| Description | A ratiometric fluorescent assay that measures mitochondrial membrane potential in live cells using the lipophilic cationic carbocyanine dye JC-1 (5,5',6,6'-tetrachloro-1,1',3,3'-tetraethylbenzimidazolylcarbocyanine iodide). JC-1 forms J-aggregates emitting red fluorescence in polarized mitochondria and exists as monomers emitting green fluorescence when mitochondrial membrane potential collapses, providing a ratiometric readout. |
| Intended Use | Assessment of mitochondrial health in early apoptosis detection, mitochondrial toxicity screening, metabolic research, and evaluation of compounds affecting mitochondrial function. |
| Principle / Technology | In cells with high mitochondrial membrane potential (ΔΨm), JC-1 enters the mitochondrial matrix and accumulates to concentrations exceeding the critical aggregation threshold, forming J-aggregates with red fluorescence (Ex/Em: 585/590 nm); upon mitochondrial depolarization during early apoptosis or metabolic stress, JC-1 remains as green-fluorescent monomer (Ex/Em: 510/527 nm); the red-to-green fluorescence ratio reports ΔΨm changes independent of dye loading variations. |
| Detection Method | Fluorescence microscopy with dual-band FITC/TRITC or GFP/RFP filter sets, flow cytometry with 488 nm excitation and FL1 (green)/FL2 (red) detection, or fluorescence plate reader |
| Sample Type | Live adherent and suspension mammalian cells, isolated mitochondria preparations, stem cells, primary neurons and cardiomyocytes |
| Performance Range / Specifications | Red/green fluorescence ratio shift of 5–10 fold between polarized and depolarized mitochondria; JC-1 monomer (green) Ex/Em: 510/527 nm; JC-1 J-aggregate (red) Ex/Em: 585/590 nm |
| Sensitivity / LOD | Detection of mitochondrial depolarization in <5% of cell population by flow cytometry; discernible red-to-green shift induced by 1 μM FCCP (uncoupler) within 15 minutes |
| Specificity | JC-1 accumulation and aggregation are potential-dependent and reversible; monomer-to-aggregate transition is specific to mitochondrial inner membrane potential; CCCP or FCCP treatment abolishes red J-aggregate fluorescence as a positive depolarization control |
| Reaction Conditions / Protocol | Prepare JC-1 working solution (dilute DMSO stock to 10 μg/mL in culture medium or assay buffer); replace cell medium with JC-1 working solution; incubate 15–30 minutes at 37°C with 5% CO2; wash twice with PBS or assay buffer; analyze by fluorescence microscopy or flow cytometry; include FCCP/CCCP-treated cells as depolarization positive control |
| Components / Formulation | JC-1 dye (lyophilized, 5 mg or 1 mg vials or 10 mg/mL DMSO stock), 10× Assay Buffer (HEPES-buffered saline), CCCP mitochondrial uncoupler (10 mM DMSO stock) as positive control, detailed protocol for microscopy and flow cytometry applications |
| Storage Conditions | Lyophilized JC-1: -20°C protected from light and moisture for 12 months; JC-1 DMSO stock (10 mg/mL): -20°C protected from light, desiccated, stable 6 months; working solution: prepare fresh and protect from light |
| Shelf Life | 12 months from date of manufacture for lyophilized dye |
| Package Specifications | 100 tests (microscopy), 100 tests (flow cytometry), bulk sizes available |
| Product Form | Lyophilized dye or concentrated DMSO stock with separate assay buffer and control reagents |
| Quality Control | Each lot tested using HeLa cells with and without 50 μM FCCP treatment; J-aggregate/monomer ratio shift ≥5-fold confirmed; lot-to-lot fluorescence intensity verified within 15% |
| Key Features | Ratiometric measurement eliminates variations from unequal dye loading or cell number; detects early mitochondrial events preceding caspase activation; compatible with multi-well plate reader for high-throughput screening; CCCP positive control included for signal range validation |
For research use only, not for clinical use.
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