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| Product Name | Mitochondrial Complex I Activity Microplate Assay Kit |
| Catalog No. | CMTR-HMM-0066 |
| Description | A colorimetric microplate assay for measuring NADH dehydrogenase activity in isolated mitochondria and tissue homogenates. The assay monitors the oxidation of NADH to NAD+ coupled to the reduction of a synthetic electron acceptor dye that produces a colored product. Complex I activity is defined as the rotenone-sensitive portion of the total NADH oxidation rate, with rotenone included as a selective Complex I inhibitor control. |
| Intended Use | Quantitative measurement of mitochondrial respiratory chain Complex I enzymatic activity for mitochondrial function assessment. |
| Principle / Technology | NADH oxidation coupled to dye reduction with rotenone-sensitive activity subtraction |
| Detection Method | Colorimetric (absorbance at 600 nm, kinetic mode) |
| Sample Type | Isolated mitochondria, tissue homogenates (muscle, liver, brain, heart), permeabilized cells |
| Performance Range / Specifications | Linear rate measurement: 30 min kinetic read at 30 °C; activity expressed as nmol NADH oxidized/min/mg protein |
| Sensitivity / LOD | Detection limit: 0.5 nmol NADH per min in a 200 µL reaction |
| Specificity | Rotenone-sensitive activity defines Complex I-specific contribution; rotenone-insensitive component reflects non-specific NADH oxidation |
| Reaction Conditions / Protocol | Add mitochondrial sample to assay buffer containing NADH and electron acceptor dye; record absorbance kinetics with and without rotenone for 30 min; calculate rotenone-sensitive rate |
| Components / Formulation | Assay buffer (potassium phosphate, pH 7.5, with BSA), NADH substrate, electron acceptor dye, rotenone inhibitor solution, mitochondrial extraction buffer |
| Storage Conditions | NADH and dye: –20 °C; buffers: 2–8 °C; rotenone: –20 °C protected from light |
| Shelf Life | 6 months for NADH and dye at –20 °C |
| Package Specifications | 100 tests |
| Product Form | Liquid reagents and lyophilized enzyme substrates |
| Quality Control | Functional validation with bovine heart mitochondrial extract; rotenone inhibits ≥80% of total NADH oxidation |
| Key Features | Rotenone control well built into assay protocol provides per-sample Complex I specificity determination |
| Purity | Reagent grade; fluorescent probes ≥95% pure by HPLC |
| Concentration | As specified per kit format; probe stock solutions in DMSO as specified |
| Activity / Unit Definition | Standard curve linearity R² ≥0.99 per lot; dynamic range as specified |
| Molecular Weight | Varies by probe and buffer component |
| Source / Origin | Synthetic fluorescent probes; recombinant enzymes; purified metabolite standards |
| pH Range / Optimal pH | pH 7.2–7.4 for cell-based assays; pH varies for biochemical metabolite detection |
| Shipping Conditions | Cold packs or dry ice for probes and enzymes; protect from light |
| Expiration Date / Stability | 6–12 months at recommended storage temperature |
| Regulatory / Compliance | Research use; manufactured under ISO 9001 |
| Compatibility | Compatible with fluorescence microplate readers, luminometers, and spectrophotometers |
| Recommended Buffer System | Assay-specific buffer optimized per product protocol |
| Application Notes / Precautions | Normalize signal to cell number or protein content; avoid probe photobleaching; include appropriate antioxidant controls for ROS assays |
| Batch-to-Batch Consistency | Standard curve parameters within ±15% of reference lot |
For research use only, not for clinical use.
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