Technical Reference #1850

1850.

Binding Site on the Transferrin Receptor for the Parvovirus Capsid and Effects of Altered Affinity on Cell Uptake and Infection Laura B. Goodman; Sangbom M. Lyi; Natalie C. Johnson; Javier O. Cifuente; Susan L. Hafenstein; and Colin R. Parrish, Cornell University College of Veterinary Medicine, Journal of Virology, 84(1850), (2010)
Link To Paper

Materials & Methods:
Fluorescence microscopy. Stable TRVb cell lines expressing variant TfRs were seeded at a density of 1 104 cells/cm2 in coverslip dishes (MatTek Ashland MA) and then used for experiments 2 days later. To detect the early stages of entry the TfR cell lines were incubated with Alexa488– or Alexa594–CPV-2 capsids or with Texas Red- or Alexa488-labeled Tf at 37°C in phenol-red free DMEM. The binding and distribution of the virus or Tf were observed within 15 min of incubation and then the cells were washed and incubated at 37°C for various times (17). Cells were also fixed with 4% paraformaldehyde after 15 30 or 60 min of incubation at 37°C. Images were collected with a Hamamatsu OrcaER camera with different labels collected as separate channels and then analyzed using the SimplePCI software (Hamamatsu Sewickley PA).

Microscopic Technique
Fluorescence Microscopy

Cell Type(s)
CHO cells