Expression of immunogenic structural proteins of XXXX in vitro assessed using immunofluorescence.

@entity7136 @entity28868 @entity281
@entity28868 (CyHV-3), also called @entity28868 (KHV), is the aetiological agent of a fatal disease in @entity7136 and @entity7136 ( @entity7136 L.), referred to as @entity28868 disease. The virus contains at least 40 structural proteins, of which few have been characterised with respect to their immunogenicity. Indirect immunofluorescence assays (IFAs) using two epitope-specific monoclonal antibodies (MAbs) were used to examine the expression kinetics of two potentially immunogenic and diagnostically relevant viral antigens, an envelope glycoprotein and a capsid-associated protein. The rate of expression of these antigens was determined following a time-course of @entity281 in two CyHV-3 susceptible cell lines. The results were quantified using an IFA, performed in microtitre plates, and image analysis was used to analyse confocal micrographs, enabling measurement of differential virus-associated fluorescence and nucleus-associated fluorescence from stacks of captured scans. An 8-tenfold increase in capsid-associated protein expression was observed during the first 5  days post- @entity281 compared to a  <=2-fold increase in glycoprotein expression. A dominant protein of ~100  kDa reacted with the capsid-associated MAb (20F10) in western blot analysis. This band was also recognised by sera obtained from @entity7136 infected with CyHV-3, indicating that this capsid-associated protein is produced in abundance during @entity281 in vitro and is immunogenic to @entity7136 . Mass spectrometry carried out on this protein identified it as a previously uncharacterised product of open reading frame 84. This abundantly expressed and immunogenic capsid-associated antigen may be a useful candidate for KHV serological diagnostics.
