Technical Reference #1761

1761.

Early Steps in Cell Infection by Parvoviruses: Host-Specific Differences in Cell Receptor Binding but Similar Endosomal Trafficking Carole E. Harbison; Sangbom Michael Lyi; Wendy S. Weichert; Colin R. Parrish, Cornell University College of Veterinary Medicine, Journal of Virology, 83(1761), (2009)
Link To Paper

Materials & Methods:
Cells and viruses. Feline CRFK and NLFK cells and canine A72 and Cf2Th cells were grown in a 1:1 mixture of McCoy’s 5A and Liebovitz L15 media with 5% fetal bovine serum. Chinese hamster ovary-derived cells lacking the hamster TfR (TRVb cells) (39) were grown in Ham’s F12 medium containing 5% fetal bovine serum. Viruses were derived from infectious plasmid clones of FPV (FPV-b) CPV-2 (CPV-d) and CPV-2b (CPV-39) strains (27 47). Plasmids were transfected into NLFK cells and the viruses recovered were titrated using 50% tissue culture infective dose (TCID50) assays (46). Virus capsids were concentrated by polyethylene glycol precipitation followed by sucrose gradient centrifugation and then dialyzed against either phosphate-buffered saline (PBS) or 20 mM Tris-HCl (pH 7.5) and stored at 4°C (1 40).

Microscopic Technique
Fluorescence Microscopy

Cell Type(s)
CRFK & NLFK cells; A72 & Cf2Th cells; CHO cells