Wednesday, 17 November 2004
D0461

Evaluation of commercial protein transduction reagents for use in an insect cell line

Kipper E. Van Meter, KVanMeter@atsu.edu and Melissa K. Stuart, MStuart@atsu.edu. Kirksville College of Osteopathic Medicine - A.T. Still University, Dept. of Microbiology/Immunology, 800 W. Jefferson St, Kirksville, MO

Protein transduction is the process by which peptides or proteins cross the cellular plasma membrane. Direct delivery of proteins into cells by-passes the need for DNA cloning and gene expression to evaluate a protein’s biological effect. The technique has been used for a variety of applications, including elucidation of signal transduction pathways, blocking of transcription factors, and structure/function analyses on the cytoplasmic domains of receptor molecules. Several commercial protein transduction reagents are available to facilitate the introduction of peptides and proteins into mammalian cells, but none of these have been reported useful in insect cell lines. In this study, we evaluated eight different protein transduction reagents from four commercial suppliers for the ability to transduce fluorescently-labeled calf histone H1 and/or goat antibody into Sf21 cells. Four of the reagents were proprietary non-lipid formulations. All of these appeared toxic for the insect cells, since the cells that fluoresced--and thus were presumably transduced--lacked plasma membrane integrity when examined by light microscopy or Trypan blue exclusion, while cells that did not fluoresce remained healthy. Sf21 cells incubated with any of the four cationic lipid-based reagents maintained good viability, but the reagents varied in their ability to fuse with the insect cell membrane and/or form complexes with the labeled proteins. Currently, the MegaFectin product line from Qbiogene, which is marketed as a DNA transduction reagent, shows the greatest promise for successful protein transduction into Sf21 cells, although further studies are needed to optimize the transduction protocol.


Species 1: Lepidoptera Noctuidae Spodoptera frugiperda (fall armyworm)
Keywords: protein transduction, Sf21 cells

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