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A FACS-measured immunofluorescence assay was developed for the detection of antibodies directed against the hemagglutinin (H) and fusion (F) glycoproteins of measles virus (MV). Human melanoma cell lines transfected with either the MV H or F genes, which showed a high surface expression of the respective proteins in their native conformation, were used as target cells. The cells were incubated with diluted plasma samples, and stained subsequently with FITC-conjugated secondary antibodies. The FACS-measured fluorescence signals correlated directly with the amount of specific immunoglobulins over a wide concentration range. The use of different conjugates enabled the separate detection of MV-specific IgG, IgM, IgA and IgG subclasses, with relatively low backgrounds. Hemagglutinin-specific IgG, IgM and IgA fluorescence signals were shown to correlate well with MV-specific IgG ELISA titers and MV-specific IgM or IgA capture ELISA OD450-values, respectively. The polyclonal conjugates with specificity for human immunoglobulins offered sufficient cross-reactivity to detect MV-specific IgG, IgM and IgA in plasma samples of cynomolgus macaques, making this technique a useful tool for studying serological responses in vaccination and challenge experiments in non-human primate models.

Immuno affinity chromatography with virus neutralizing monoclonal antibodies, directed to the haemagglutinating protein of canine parvovirus (CPV) was used to purify and concentrate CPV from infected cell culture. The procedure was monitored by testing the respective fractions in an infectivity titration system, in an ELISA, in a haemagglutination assay and by negative contrast electron microscopy to quantify CPV or CPV antigen. The degree of purification was further estimated by testing the fractions for total protein content in a colorimetric method, for bovine serum albumin content in an ELISA and by SDS-PAGE. Over 99% of the contaminating proteins proved to be removed, and 20% or 70-90% of infectious CPV or CPV antigen, respectively, was recovered.