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• Viral RNA titration in Chikungunya inoculated tissue samples

Viral RNA titration in Chikungunya inoculated tissue samples

Instrument: Precellys® 24 Dual

Sample: Splenic, hepatic, muscular, and joint tissues from cynomolgus macaques inoculated with CHIKV (30 mg /~100mg)

Protocol:

• Precellys® kit: 03961-1-002 (ceramic beads 2.8mm) for smooth tissues and 03961-1-001 (metals beads 2.8mm) for joint and muscle.
• Add samples: Splenic, hepatic, muscular, and joint tissues from cynomolgus macaques inoculated with CHIKV (30 mg /~100mg).
• Add buffer: 1 ml lysis buffer for Viral RNA extraction or 1 ml DMEM supplemented with 10% FCS for Viral titration
• Run the Precellys at 5000rpm, 2 x 20sec or 2x10sec 
• Tissue viral RNA extraction: Nucleospin 96 RNA kit, Relative quantitative RT-PCR simultaneously with CHIKV and GAPDH primers and probes.

Results:

Determination of viral titers (TCID50/g): on mammalian BHK-21 cell lines based on their TCID50 using 4–5 replicates.