Comparison of indirect and direct immunofluorescence for the detection of Respiratory Syncytial Virus

Abstract

The respiratory syncytial virus (RSV) is the major cause of respiratory infection in children, with bronchopneumonia and bronchiolitis. In the adult the clinical manifestation is lighter than in infant, however, in the case of immunocompromised patients (transplant recipients or patients in immunosuppressive treatment) infection can lead to death. Immunofluorescence is a diagnostic procedures which can detect RSV infection and the presence of viable virus in the biological sample of the patient. In this study, indirect and direct immunofluorescence techniques using monoclonal antibodies directed to RSV have been compared. For the comparison HEP2 shell vials, that are susceptible to RSV, were used.These cells were infected with different virus titres, ranging from 102 to 10-3 TCID50. Subsequently, different primary or Fluorescein isothiocyanate antibody dilutions were tested (1:40, 1:80, 1:160), for indirect and direct immunofluorescence evaluation, respectively, at three incubation periods after infection (48h, 72h, 96h). Indirect immunofluorescence showed a sensitivity of 101 TCID50, with a cells positive detection even at lower dilutions up to 10-1 TCID50 at 72 hours. For direct immunofluorescence, the same sensitivity was observed with 1:40 antibody dilution at 72h, with a detection limit of 10-1 TCID50. Indirect immunofluorescence showed optimal RSV detection at 72h after infection with 1:80 primary antibody dilution, with a sensitivity of 101 TCID50. An equivalent sensitivity has been observed with direct immunofluorescence at 72h, 1:40 antibody dilution, thus representing an advantage in terms of time.