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| Product Name | Nocodazole Cell Cycle Synchronization Reagent |
| Catalog No. | CCAT-HMM-0054 |
| Description | A high-purity cell-permeable microtubule polymerization inhibitor that arrests cells at the G2/M phase boundary by disrupting mitotic spindle formation. Nocodazole synchronizes cycling cells at prometaphase by interfering with microtubule dynamics and activating the spindle assembly checkpoint. |
| Intended Use | Cell cycle synchronization for biochemical analysis of cell cycle-regulated proteins, preparation of mitotic cell populations for chromosome analysis, and study of mitotic checkpoint signaling mechanisms. |
| Principle / Technology | Nocodazole (methyl [5-(2-thienylcarbonyl)-1H-benzimidazol-2-yl]carbamate) binds to β-tubulin with high affinity and prevents the assembly of tubulin dimers into microtubules; inhibition of spindle microtubule polymerization activates the spindle assembly checkpoint (SAC), blocking the anaphase-promoting complex/cyclosome (APC/C) and arresting cells at the G2/M transition; the effect is reversible upon compound washout, allowing synchronized release into subsequent cell cycle phases. |
| Detection Method | Flow cytometric DNA content analysis (propidium iodide staining) to verify G2/M accumulation; phospho-histone H3 (Ser10) immunostaining for mitotic index; time-lapse microscopy for mitotic progression monitoring |
| Sample Type | Proliferating adherent and suspension mammalian cell lines; typically HeLa, U2OS, HEK293, CHO, and other rapidly dividing cell types |
| Performance Range / Specifications | Optimal working concentration: 40–100 ng/mL (0.13–0.33 μM) depending on cell type; treatment duration: 12–18 hours for complete G2/M arrest; typical mitotic index yield: 70–95% of cell population; reversible release within 30–60 minutes of washout |
| Sensitivity / LOD | Achieves >80% G2/M arrest in HeLa cells at 50 ng/mL with 16-hour treatment; synchronized release yields >60% cells progressing synchronously through G1 phase within 2 hours |
| Specificity | Nocodazole specifically binds β-tubulin with a Kd of approximately 0.2 μM; minimal binding to actin or intermediate filaments; does not interfere with nucleic acid synthesis; microtubule depolymerization is microtubule-specific and calcium-independent |
| Reaction Conditions / Protocol | Prepare 10 mg/mL nocodazole stock in DMSO; add to culture medium at recommended concentration (typically 50 ng/mL final with ≤0.1% DMSO); incubate cells for 12–18 hours at 37°C with 5% CO2; mitotic cells can be harvested by gentle shake-off due to rounded morphology and weakened attachment; to release, wash cells twice with warm PBS and add fresh medium; harvest at desired time points for cell cycle analysis |
| Components / Formulation | Nocodazole (≥99% purity by HPLC) supplied as lyophilized powder; molecular weight 301.3 g/mol; soluble in DMSO at ≥20 mg/mL; recommended to prepare 10 mg/mL stock in anhydrous DMSO and store in single-use aliquots |
| Storage Conditions | Lyophilized powder: -20°C protected from light and moisture, stable 36 months; DMSO stock (10 mg/mL): -20°C protected from light and moisture, stable 6 months |
| Shelf Life | 36 months from date of manufacture for lyophilized powder |
| Package Specifications | 10 mg, 25 mg, 50 mg, 100 mg lyophilized powder |
| Product Form | Lyophilized powder requiring dissolution in DMSO |
| Quality Control | Each lot analyzed by HPLC for purity (≥99% specification); biological activity confirmed in HeLa cell cycle arrest assay with >80% G2/M accumulation at 50 ng/mL; residual solvent tested; heavy metals within ICH Q3D limits |
| Key Features | Most widely used G2/M synchronizing agent in cell cycle research; rapid and fully reversible activity; effective across a broad range of mammalian species and cell types; compatible with subsequent proteomic, genomic, and biochemical analyses; mitotic shake-off enables enrichment without trypsinization |
For research use only, not for clinical use.
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