INVESTIGADORES
PEREZ FILGUEIRA Daniel Mariano
artículos
Título:
Optimization and validation of recombinant serological tests for African swine fever diagnosis based on the p30 protein produced in Trichoplusia ni larvae.
Autor/es:
PÉREZ FILGUEIRA, DANIEL MARIANO; GONZÁLEZ-CAMACHO, F.; GALLARDO, C.; RESINO-TALAVAN, P.; BLANCO, E.; GOMEZ-CASADO, E.; ALONSO, C.; ESCRIBANO, J. A. M.
Revista:
JOURNAL OF CLINICAL MICROBIOLOGY
Editorial:
American Society for Microbiology (ASM)
Referencias:
Año: 2006 vol. 44 p. 3114 - 3121
ISSN:
0095-1137
Resumen:
We describe the validation of an enzyme-linked immunosorbent assay (ELISA) and confirmatory immunoblotting
assays based on a recombinant p30 protein (p30r) produced in insect larvae using a baculovirus
vector. Such validation included the following: (i) the scaling up and standardization of p30r production and
the associated immunoassays, (ii) a broad immunological analysis using a large number of samples (a total of
672) from Spain and different African locations, and (iii) the detection of the ASF virus (ASFV)-antibody
responses at different times after experimental infection. Yields of p30r reached up to 15% of the total protein
recovered from the infected larvae at 3 days postinfection. Serological analysis of samples collected in Spain
revealed that the p30r-based ELISA presented similar sensitivity to and higher specificity than the conventional
Office International des Epizooties-approved ASFV ELISA. Moreover, the p30r ELISA was more
sensitive than the conventional ELISA test in detecting ASFV-specific antibodies in experimentally infected
animals at early times postinfection. Both the recombinant and conventional ELISAs presented variable rates
of sensitivity and specificity with African samples, apparently related to their geographical origin. Comparative
analyses performed on the sequences, predicted structures, and antigenicities of p30 proteins from different
Spanish and African isolates suggested that variability among isolates might correlate with changes in
antigenicity, thus affecting detection by the p30r ELISA. Our estimations indicate that more than 40,000 ELISA
determinations and 2,000 confirmatory immunoblotting tests can be performed with the p30r protein obtained
from a single infected larva, making this a feasible and inexpensive strategy for production of serological tests
with application in developing countries.