Application of real-time fluorescent PCR for quantitative assessment of Neospora caninum infections in organotypic slice cultures of rat central nervous system tissue.
The previously described Nc5-specific PCR test for the diagnosis of Neospora caninum infections was used to develop a quantitative PCR assay which allows the determination of infection intensities within different experimental and diagnostic sample groups. The quantitative PCR was performed by using a dual fluorescent hybridization probe system and the LightCycler Instrument for online detection of amplified DNA. This assay was successfully applied for demonstrating the parasite proliferation kinetics in organotypic slice cultures of rat brain which were infected in vitro with N. caninum tachyzoites. This PCR-based method of parasite quantitation with organotypic brain tissue samples can be regarded as a novel ex vivo approach for exploring different aspects of cerebral N. caninum infection.
Müller
Norbert
N
Institute of Parasitology, University of Berne, Länggass-Strasse 122, 3012 Berne, Switzerland. nmueller@ipa.unibe.ch
Vonlaufen
Nathalie
N
Gianinazzi
Christian
C
Leib
Stephen L
SL
Hemphill
Andrew
A
D000818:Animals; D001921:Brain / Q000469:parasitology*; D002540:Cerebral Cortex / Q000166:cytology; D003048:Coccidiosis / Q000469:parasitology*; D016054:DNA, Protozoan / Q000032:analysis; D007150:Immunohistochemistry; D018688:Neospora / Q000235:genetics / Q000254:growth & development / Q000302:isolation & purification*; D009924:Organ Culture Techniques; D016133:Polymerase Chain Reaction / Q000379:methods*; D051381:Rats
D016054:DNA, Protozoan