Methods for assessing feline immunodeficiency virus infection, infectivity and purification

Abstract

Infection of cats with the feline immunodeficiency virus (FIV) recapitulates many aspects of infection of humans with HIV, including highly activated but ineffectual immune responses. Infected hosts remain seropositive for life, and detection of antibodies is the mainstay of diagnosis. However, to quantify virus for research or prognosis, viral proteins, nucleic acids or enzymes, are typically measured by ELISA, PCR or activity, respectively. While such assays are in wide use, they do not distinguish whole, infectious viral particles from defective or disrupted viruses. Titers of infectious viral particles may be estimated from tissue culture infectious doses or by enumerating cell-associated viral proteins, viral transcriptional activity or formation of syncytia. To analyze the viral proteome and the incorporation of host components into viral envelopes, pure lentiviral preparations are required. Methods for purifying lentiviruses include ultracentrifugation to separate particles by size, mass and/or density; chromatography to separate particles by charge, affinity or size; and additional removal of extraviral proteins and exosomes through subtilisin digestion or immunoaffinity. This article reviews advantages and disadvantages of different approaches to purification of lentiviruses with special reference to suitability for FIV, and highlights effects of purification on immune responses and immune assays.