1. An Introduction to the Virology Laboratory A History of Virology Virology Laboratory design and equipment used Definition of Biosafety Levels in Virology Laboratories (BSL1-4) Virology Laboratory Codes of Practices Personal Protective Equipment in the Virology Laboratory (BSL1-4) Storage and transportation of viruses 2. Cell Culture in Virology Aseptic Technique and working in an MSCII Preparation of cell culture media Preparation of continuous cell culture Preparation of primary cell cultures Maintenance of cell cultures Cryopreservation of cells Quantification of cells Transformation of bacteria for virus cDNA clone production Mini and Midi Prep purification from bacterial cultures Electroporation of cells with virus nucleic acid 3. Cell based virology protocols Virus propagation Cell culture Eggs Animal models (IN/IC/IP/SC) Arthropods Virus Harvest Cell culture Animal tissues & blood Eggs Environmental samples Arthropods Human clinical samples Arthropod saliva extraction Plaque Assay ? infectious virus particle titration Plaque purification Tissue Culture Infectious Dose (TCID50) - virus titration Egg Infectious Dose(EID50) - - virus titration Plaque reduction Neutralisation Test (PRNT) ( see serology-based protocol) Viral burden by measurement ATP depletion 4. Serology-based virology protocols Haemagglutination Assay (HA) Haemagglutination Inhibition Assay (HAI) Plaque Reduction Neutralisation Test (PRNT) Immunofluorescence slide preparation (IFA) IgG / IgM Indirect ELISA OgG / IgM Direct ELISA Interferon ELISA Flow cytometry 5. Molecular virology protocols Nucleic acid extraction (manual / automated / high throughput) Primer design Probe design Thermocycling program design Polymerase Chain Reaction (PCR) Reverse Transcription-PCR Real-Time PCR / RT-PCR PCR Platforms (ABI, Roche, BioRad) Realtime PCR platforms (ABI, Roche, BioRad) Gel Electrophoresis Preparation of PCR amplicons for Sanger sequencing Gel purification of Nucleic acid Terminal Dye labeling and purification Preparation of PCR amplicons for NGS Loop mediated isothermal amplification (LAMP) Microarray slide preparation 6. Cloning Virus expression in cDNA clones Reverse transcription for electroporation Protein expression in virus vectors Restriction enzymes 7. Visualization of Virus Preparation of samples for electron microscopy Understanding cytopathic effect (CPE) of infected cellc 8.
Novel protocols in virology 3D Digital PCR ViroCyt Next Generation Sequencing (NGS) Immunochromatographic assays &Lateral Flow devices Virus propagation in Plants 9. Annexes Hazard group and containment list for each virus Geographical distribution matrix for viruses Worked examples of calculations for each protocol List of laboratory equipment and reagents used in each protocol List of labs with expertise.