LipoD293™ In Vitro DNA Transfection Reagent

Description:
Leveraging our innovative and proprietary lipid conjugation technology, LipoD293™ (Ver. II) is specifically designed and formulated with proprietary enhancers for highly efficient DNA transfection in HEK293 cells and other mammalian cell types (Figure 1). As a second-generation liposome-based DNA transfection reagent, LipoD293™ (Ver. II) delivers exceptionally high transfection efficiency in HEK293-derived cells as well as many other mammalian cells, while maintaining low cytotoxicity.

Compared with the previous formulation, LipoD293™ (Ver. II) incorporates a refined chemistry that results in a 3–4 fold improvement in gene delivery efficiency. One 1.0 mL vial of LipoD293™ (Ver. II) Reagent is sufficient for approximately 666 transfections in 24-well plates or 333 transfections in 6-well plates.

Figure 1. Schematic illustration of enhanced gene delivery mediated by LipoD293™ (Ver. II).

Features:
– Preferred choice for hard-to-transfect cells
– Enables exceptionally high viral titers
– Highly effective for transfection of very large DNA constructs (up to 180 kb)
– Performs equally well in suspension HEK293 cells (e.g., 293F, 293H)
– Supports high-level recombinant protein expression
– Transfection efficiency is enhanced in the presence of serum and antibiotics in 293 cells
– Exceptional performance for both single-DNA transfection and multi-DNA co-transfection
– Highly cost-effective transfection solution

Storage Condition:
Store at 4 °C. When stored under recommended conditions, the product is stable for 12 months or longer.

Comparisons of Transfection Efficiency of LipoD293™ DNA In Vitro Transfection Reagent (Ver. II) with Brand Name Products

Comparison of transfection efficiency of LipoD293™ reagent (Ver. II) vs. lipofectamine 2000 (L2K) and Fugene HD on HepG2 cells.
Right Panel: Comparison of transfection efficiency of LipoD293 (Ver. II) with Lipofectamine 2000 (L2K), and Fugene HD on HepG2 cells. GFP DNA (pEGFP-N3) was transfected with different transfection reagents per the manufacturer’s protocols to HepG2 cell (cultured on collagen-pretreated dishes). GFP-positive cell (%) and fluorescence intensity were detected by passing through FACS 48 hours post-transfection
Left Panel: presence of serum and antibiotics enhances LipoD293 (Ver. II) efficiency on HepG2 cells. HepG2 cell (grown on collagen-treated dishes) was transfected with three different conditions——-serum and antibiotics free, presence of 10% serum and antibiotics, followed by removal 5 hours post transfection, and presence of 10% serum and antibiotics without removal 5 hours post transfection.

Comparison of transfection efficiency of LipoD293™ reagent (Ver. II) vs. lipofectamine 2000 (L2K), TransIT and Fugene 6 on CHO cells.
Right Panel: Comparison of transfection efficiency of LipoD293 (Ver. II) with Lipofecatmine 2000 (L2K), TransIT and Fugene 6 on CHO cells. DNAs encoding Renilla luciferase (phRL-CMV) and GFP (pEGFP-N3) were transfected with different DNA transfection reagent per the manufacturer’s protocols. Renilla luciferase activity and GFP fluorescence were detected with Renilla Assay System and a Nikon Eclipse fluorescent microscope, respectively, 24 hours post-transfection.
Left Panel: Comparison of price ($/1.0 ml vial) of LipoD293 versus those of Lipofecatmine 2000 (L2K), TransIT, and Fugene 6. All the prices were collected from the manufacturers’ websites.

A comparison of the transfection efficiency of LipoD293™ reagent with Lipofectamine 2000 (L2K) on a hard-to-transfect cell, primary rat aortic smooth muscle cells. The rat aortic smooth muscle cells were prepared and transfected with pEGFP-N3 by LipoD293™ reagent (left panel) and Lipofectamine 2000 (L2K, right panel), respectively per the manufacturers’ protocols. The transfection efficiency was evaluated by detecting GFP fluorescence with a Nikon Eclipse 2000 microscope 24 hours post-transfection. The above pictures were kindly provided by Dr. Nickolai Dulin of the Section of Pulmonary and Critical Care, University of Chicago.

A comparison of the transfection efficiency of LipoD293™ reagent with Fugene HD on hard-to-transfect cells, LNCap cells. The LNCap cells were grown according to ATCC recommended procedures and co-transfected with pBabe-hygro-SSeCKs (1.5 ug) and pEGFP-N3 (0.5 ug) per well (6-well plate), LipoD293™ reagent (left panel) and Fugene HD (right panel), respectively per the manufacturers’ protocols. The transfection efficiency was evaluated by detecting GFP fluorescence with a Nikon Eclipse 2000 microscope 24 hours post-transfection. The above pictures were kindly provided by Dr. Lyn Gao of Roswell Park Cancer Institute.

Two examples showing the exceptional efficiency of LipoD293™ reagent on hard-to-transfect cells like HepG2 and SaoS-2 cells. HepG2 and SaoS-2 cells in 95% confluency were transfected with pEGFP-N3 and pSV-β-galactosidase DNAs, respectively in the presence of serum/antibiotics. The efficiency was checked 48 hours post-transfection by Zeiss 510 Confocal Microscopy and β-galactosidase staining kit, respectively.

A comparison of the transfection efficiency of LipoD293™ reagent with 293fectin on HEK293 cells. HEK293 cells transfected with the pEGFP-C1 plasmid using LipoD293™ In Vitro DNA Transfection Reagent (Ver. II) (upper panel) and the most popular brand product 293fectin™ of Invitrogen (lower panel). The cells were visualized by Nikon Eclipse Fluorescence microscope with DIC phase imaging (left panel) and FITC imaging (right panel) 24 hours post-transfection.

A comparison of LipoD293™ reagent vs. 293fectin, Xfect, and Fugene 6 transfection reagents on protein production with suspension 293F cells. 30 mL of 293F cell cultured in standard culture medium was transfected with pEGFP-6xHis plasmid using LipoD293™ In Vitro DNA Transfection Reagent (Ver. II) (20 ug plasmid DNA), 293Fectin (30 ug plasmid DNA), Xfect (30 ug plasmid DNA) and Fugene 6 (30 ug plasmid DNA) per manufacturers’ standard transfection protocols. GFP fluorescence was visualized 48 hours post-transfection (left panel) with A for LipoD293, B for 293fectin, C for Xfect, and D for Fugene 6. The 6xHis tagged GFP protein was then purified via Ni-NTA affinity column. 5 μL of 1st elution fraction was resolved on SDS-PAGE, followed by Coomassie Brilliant Blue staining (right upper panel E) with lane 1 for LipoD293, lane 2 for protein marker, lane 3 for Xfect, lane 4 for 293fectin, and lane 5 for Fugene 6. The protein yield was quantified via a spectrometer (right lower panel F).

A comparison of LipoD293™ (Ver. II) and Lipofecatmine LTX (LTX) on the generation of Lentivirus (LV). Three cDNAs were co-transfected with LipoD293™ (Ver. II) and Lipofectamine LTX (LTX) into 293FT cells. A GFP vector, pHR-SIN-cppt-CMVEWP, was used to determine the titer of LV. 1×105 293F cells per well were plated into a 24-well plate followed by the addition of different amounts of the vector supernatant, 1 microliter (upper panel) and 10 microliters (lower panel), respectively. 5 days later, the cells were passed with FACS. The numbers in the upper right corner indicate the percentage of transduced cells. The titers of LV generated with LipoD293™ and L2K were quantified to be 8×10^6 and 3×10^6 tu/ml, respectively

Technical Information & Datasheet:
– A General Protocol for Transfecting Mammalian Cells
– A Short Protocol for Advanced Users
– A Protocol for Packaging Lentivirus in HEK293T Cells
– A Protocol for Packaging AAV in HEK293T Cells
– A Protocol for Packaging Adenovirus in HEK293A Cells
– A Protocol for Transfecting Suspension 293 and CHO Cells
– Technical Note & Transfection Tips