TransIT-X2® Dynamic Delivery System Outperforms Lipofectamine® Reagents. A549 (A) or MDCK (B) cells were transfected with luciferase encoding plasmid DNA using either TransIT-X2® (Mirus Bio), Lipofectamine® 2000 (Thermo Fisher Scientific) or Lipofectamine® 3000 (Thermo Fisher Scientific) for 24 hours at indicated reagent-to-DNA ratios or reagent-to-P3000™-to-DNA ratio. Transfection was measured by luciferase activity using a conventional assay. Cytotoxicity was assessed by quantifying the LDH released from the cytosol of damaged cells compared to cells alone. Higher transgene expression and lower cytotoxicity was observed in cells transfected with TransIT-X2® at optimal ratios compared to cells transfected with Lipofectamine® 2000 or Lipofectamine® 3000.

Visualization of High GFP Expression Using TransIT-X2® Dynamic Delivery System. TransIT-X2® Dynamic Delivery System was used to transfect plasmid DNA encoding EGFP into A549, CHO-K1, HepG2, LNCaP, MDCK, PC12, primary human mammary epithelial cells (HMEC) and normal human dermal fibroblasts (NHDF). Transfections were performed in 35 mm MatTek dishes using 4-8 µl of TransIT-X2® to deliver 2 µg of DNA. Images (32X) were captured at 48 hours post-transfection using a Zeiss Axiovert S100 inverted fluorescence microscope. * indicates primary cell types

TransIT-X2® Outperforms Lipofectamine® for RNP Delivery. Ribonucleoprotein (RNP) complexes composed of PPIB (cyclophilin B) targeting 2-part gRNA (IDT) and Cas9 protein (PNA Bio) were delivered into HEK293T/17 and U2OS cells using TransIT-X2® Dynamic Delivery System (1 µl/well, Mirus Bio) or Lipofectamine® CRISPRMAX™ (1.5 µl/well and 1 µl/well of Lipofectamine® Cas9 Plus™ Reagent, ThermoFisher) or Lipofectamine® RNAiMAX (1.5 µl/well, ThermoFisher) or Lipofectamine® 3000 (1.5 µl/well and 1 µl/well of P3000™ Reagent, ThermoFisher) in a 24-well format according to the manufacturers’ protocol. Varying levels of gRNA (6 nM or 12 nM) were tested with 6 nM Cas9 protein (PNA Bio). A T7E1 mismatch detection assay was used to measure cleavage efficiency at 48 hours post-transfection.