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| Product Name | Hot-Start Taq DNA Polymerase (Antibody-Mediated) |
| Catalog No. | IEC-HMM-0047 |
| Description | A thermostable DNA polymerase formulated with a blocking monoclonal antibody that inhibits enzyme activity below 70 °C. The antibody dissociates upon initial heat denaturation, releasing fully active polymerase for specific target amplification. This hot-start mechanism effectively suppresses primer-dimer formation and non-specific product generation during reaction setup. |
| Intended Use | Designed for hot-start PCR, quantitative real-time PCR (qPCR), high-throughput genotyping, and any thermal cycling application where non-specific amplification must be minimized. |
| Principle / Technology | Monoclonal antibody binds to the polymerase active site at ambient temperature, sterically blocking substrate access. Upon heating above ~70 °C during the initial denaturation step, the antibody irreversibly denatures and the polymerase regains full catalytic activity. |
| Detection Method | DNA synthesis activity measured by incorporation of radiolabeled nucleotides into acid-insoluble fraction using activated calf thymus DNA as template-primer substrate at 74 °C. |
| Sample Type | Purified genomic DNA, plasmid DNA, cDNA, bacterial colonies, and synthetic oligonucleotide templates. |
| Performance Range / Specifications | Specific activity: >= 20,000 U/mg. Extension rate: 1-2 kb/min at 72 °C. Amplicon size range: 100 bp to 5 kb. 5'-3' exonuclease activity present; no 3'-5' proofreading activity. |
| Sensitivity / LOD | Detects single-copy targets from as little as 10 pg human genomic DNA in a 25 uL reaction after 35-40 amplification cycles. |
| Specificity | Antibody-mediated hot-start reduces primer-dimer and non-specific amplification by more than 95% compared to standard Taq polymerase at room temperature setup. |
| Reaction Conditions / Protocol | Initial activation: 94-95 °C, 30 s to 2 min. Denaturation: 94-95 °C, 10-15 s. Annealing: 55-68 °C, 10-30 s. Extension: 72 °C, 15-30 s/kb. 25-40 cycles. Final extension: 72 °C, 5 min. |
| Components / Formulation | Taq DNA Polymerase (5 U/uL) in buffer containing 20 mM Tris-HCl (pH 8.0), 100 mM KCl, 0.1 mM EDTA, 1 mM DTT, 50% glycerol, and stabilizing detergents, complexed with anti-Taq monoclonal antibody. Supplied with 10x PCR Buffer and 50 mM MgCl₂. |
| Storage Conditions | Store at -20 °C to -30 °C. Avoid repeated freeze-thaw cycles; aliquot into single-use volumes when possible. |
| Shelf Life | 36 months from date of manufacture under recommended storage conditions. |
| Package Specifications | 100 U (20 uL), 500 U (100 uL), 2,500 U (500 uL), 10,000 U (2 mL). Each packaged with matching volumes of 10x PCR Buffer and 50 mM MgCl₂. |
| Product Form | Clear, colorless to pale yellow liquid solution containing glycerol as cryoprotectant. |
| Quality Control | Each production lot is tested for DNA polymerase specific activity, absence of contaminating endonuclease and exonuclease activities, RNase contamination, and functional PCR amplification using standardized control templates and primer sets. |
| Key Features | Antibody-mediated hot-start with rapid activation; no separate activation incubation needed; suitable for multiplex PCR; compatible with most real-time PCR instruments and detection chemistries. |
| Purity | >95% homogeneity by SDS-PAGE with Coomassie staining; single predominant band at ~94 kDa. |
| Concentration | 5 U/uL standard. Custom concentrations available (1, 2, 10 U/uL) upon request. |
| Activity / Unit Definition | 1 unit incorporates 10 nmol total dNTP into acid-insoluble product in 30 min at 74 °C with activated calf thymus DNA template. Specific activity >= 20,000 U/mg protein. |
| Molecular Weight | ~94 kDa (monomer, recombinant Taq polymerase). Antibody component: ~150 kDa (IgG subclass). |
| Source / Origin | Recombinant enzyme produced in Escherichia coli. Gene sequence derived from Thermus aquaticus strain YT-1. |
| pH Range / Optimal pH | Optimal activity at pH 8.3-9.0 (Tris-HCl buffer). Tolerates pH 7.5-9.5 with reduced activity. |
| Shipping Conditions | Shipped on dry ice or with sufficient cold packs to maintain temperature below -15 °C during transit. Arrival at ambient temperature for short periods (<72 h) does not impair activity. |
| Expiration Date / Stability | Retains >90% activity after 36 months at -20 °C. Stable at 4 °C for up to 2 weeks. Avoid storage at temperatures above 25 °C. |
| Regulatory / Compliance | Manufactured under ISO 9001 and ISO 13485 certified quality management systems. For research use and IVD raw material supply; additional regulatory documentation available upon request. |
| Compatibility | Compatible with DMSO (up to 10% v/v), betaine (up to 1 M), formamide (up to 3% v/v), BSA (up to 0.2 mg/mL), tetramethylammonium chloride, glycerol, and common commercial qPCR master mix buffers. |
| Recommended Buffer System | Recommended 1x reaction buffer: 20 mM Tris-HCl (pH 8.8 at 25 °C), 10 mM KCl, 10 mM (NH₄)₂SO₄, 2 mM MgSO₄, 0.1% Triton X-100. Optimal Mg²⁺: 1.5-3.5 mM final concentration. |
| Application Notes / Precautions | Prepare reactions on ice. Add enzyme last to the master mix. For GC-rich templates, add DMSO to 3-5% final. Do not vortex after adding enzyme; mix by gentle pipetting. Activation time: 30 s minimum at the chosen denaturation temperature. |
| Batch-to-Batch Consistency | Activity variation between independent production lots: <= +/- 10%. Purity profile and functional PCR performance consistent across lots. Each batch accompanied by a Certificate of Analysis. |
For research use only, not for clinical use.
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