lecture 5/6 Flashcards
ultracentrifuge
- play vital role in virology
- efficiently sediment even the smallest viruses
- used for concentration and purification of virus
rate-zonal centrifugation
-separates viral particles based on mass
isopycnic centrifugation
- boyant or equilibrium seperation
- particles separated solely based on buoyant density
other forms of virus purification and concentration
-virus purification with membrane chromatography
virus quantification
-counts the number of viruses in a specific volume to determine virus concentration
virus titer
- lowest concentration of virus that still infects cells
- number of infectious units per ml of sample
biological virus quantification tests
- depends on a virus particle initiating a successful replication cycle
- plaque assays
- pock assays
- various endpoint titration methods
physical viral quantification tests
- do not depend on any biological activity of the virus particle
- electron microscope particle counts
- hemagglutination
- immunological assays such as ELISA
- quantitative PCR assays
- flow cytometry
transmission electron microscopy (direct physical test)
-most direct method to determine the concentration of virus particles in a sample
Virus Counter 2100
- physical test
- direct counting
Assessment based on antigen concentration (physical test)
- hemagglutination assay
- single radial immunodiffusion (SRID)
Assessment based on gene expression
- quantitative polymerase chain reaction
- physical test
traditional assays/ biological assays
-plaque (circular zone of necrotic cells surrounded by viable cells in a monolayer)
Plaque forming units
- units/mL (PFU)
- measures the number of virus particles capable of forming plaques per unit volume
- ex: if a solution has PFU of 1000 pfu/ml, every ml of solution contains enough virus to form 1000 plaques
pock assay (biological assay)
- pock is necrotic area of chorioallantoic membrane of embryonated egg
- unit: pock-forming units/ml
transformation assay (biological assay)
- quantitative determination of titers of oncogenic viruses
- unit: focus-forming units/ml
quantal assay (biological assays)
-endpoint=virus dilution that affects 50% of test subjects
TCID50 (tissue culture infective dose 50)
-tissue culture infectious dose which will infect 50% of the cell monolayers challenged by the defined inoculum
Which of the following is not a physical assay for quantification of viruses?
A. electron microscopy
B. ELISA
C. pock assay
D. qPCR
C. pock assay
A PFU/mL of 300 means:
A. 300 viruses are present in 1 ml sample to cause one plaque in monolayer cell culture
B. one virus is present per ml of sample to form 300 plaques in monolayer cell culture
C. the minimum number of viruses present per ml of the sample to form 300 plaques in monolayer cell culture
B.
