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| Product Name | Endotoxin-Free Plasmid Maxiprep Kit |
| Catalog No. | NATR-HMM-0110 |
| Description | A gravity-flow anion-exchange column kit for the large-scale purification of transfection-grade plasmid DNA. The kit incorporates an endotoxin removal step that reduces bacterial endotoxin to ultra-low levels, making the purified DNA suitable for in vivo applications and sensitive mammalian cell transfections. |
| Intended Use | Large-scale isolation of high-purity, low-endotoxin plasmid DNA from 100–500 mL E. coli cultures for mammalian transfection, in vivo gene delivery, microinjection, and therapeutic-grade applications in research settings. |
| Principle / Technology | Alkaline lysis releases plasmid DNA from bacterial cells; the cleared lysate is loaded onto an anion-exchange resin that selectively binds plasmid DNA under moderate salt conditions while RNA, proteins, and metabolites flow through. A subsequent endotoxin removal wash step eliminates residual lipopolysaccharides before plasmid DNA is eluted under high-salt conditions and recovered by isopropanol precipitation. |
| Detection Method | UV spectrophotometry (A260/A280, A260/A230 ratios); agarose gel electrophoresis; LAL endotoxin assay; transfection efficiency evaluation in HEK293 or CHO cells |
| Sample Type | E. coli cultures grown in LB, TB, or 2xYT medium (100–500 mL); high-copy and low-copy plasmids; standard cloning and expression strains |
| Performance Range / Specifications | Typical yield from 250 mL overnight culture: 500–1000 μg (high-copy), 100–300 μg (low-copy); A260/A280: 1.80–1.95; endotoxin: <0.1 EU/μg plasmid DNA |
| Sensitivity / LOD | Minimum culture volume: 100 mL; effective plasmid recovery from stationary-phase cultures with OD600 > 3.0 |
| Specificity | Plasmid DNA free of genomic DNA, RNA, protein, and >99.9% reduction in bacterial endotoxin; <0.1 EU/μg DNA endotoxin level; no detectable RNase or DNase activity |
| Reaction Conditions / Protocol | Harvest bacteria from 100–500 mL culture, resuspend in 10 mL resuspension buffer, add 10 mL lysis buffer, incubate 5 minutes, add 10 mL neutralization buffer, filter lysate through included filter, load onto equilibrated column, wash with 2×30 mL medium-salt wash buffer, wash with 2×15 mL endotoxin removal buffer, elute with 15 mL high-salt elution buffer, precipitate with 0.7 volumes isopropanol, centrifuge 15,000g for 30 minutes at 4°C, wash pellet with 70% ethanol, air dry, resuspend in TE or water; total time approximately 4–5 hours |
| Components / Formulation | Resuspension buffer (50 mM Tris-HCl pH 8.0, 10 mM EDTA, 100 μg/mL RNase A), lysis buffer (200 mM NaOH, 1% SDS), neutralization buffer (3 M potassium acetate pH 5.5), equilibration buffer, wash buffer, endotoxin removal buffer, elution buffer, isopropanol, anion-exchange resin columns with reservoirs, lysate filtration units |
| Storage Conditions | RNase A-containing resuspension buffer at 2–8°C; columns, buffers, and other components at room temperature; protected from light |
| Shelf Life | 24 months for buffers and columns; resuspension buffer with RNase A stable for 6 months at 2–8°C after preparation |
| Package Specifications | 10 and 25 preparation kits (250 mL culture scale per column); all consumables included |
| Product Form | Liquid buffers; pre-packed anion-exchange resin columns; lysate filtration syringes; lyophilized RNase A |
| Quality Control | Each lot validated with pCMV-GFP plasmid: yield ≥500 μg from 250 mL DH5α; A260/A280: 1.80–1.95; >85% supercoiled; endotoxin <0.1 EU/μg; transfection efficiency ≥70% in HEK293 cells by GFP fluorescence; no residual bacterial genomic DNA by qPCR |
| Key Features | Patent-pending endotoxin removal technology achieves <0.1 EU/μg without additional clean-up steps; gravity-flow format requires no specialized equipment; concentrated plasmid solutions achievable for in vivo injections; validated for primary and stem cell transfections |
For research use only, not for clinical use.
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