Virus Quantification Flashcards
virus quantification
counts the number of viruses in a specific volume to determine the virus concentration
virus titer
lowest concentration of virus that still infects cells. Number of infectious units per ml of the sample/solution
biological quantification tests
depends on a virus particle initiating a successful replication cycle
plaque assay
pock assay
various endpoint titration methods
physical quantification tests
do not depend on any biological activity of the virus particle electron microscopic particle counts hemagglutination immunological assay, ELISA quantitative PCR assay flow cytometry
direct particle count by transmission electron microscope (TEM)
why is it not generally performed
expensive, required trained staff, changes of error are high
virus counter 2100
specialized flow cytometry, each sample is stained with two different fluorescent dyes, one for nucleic acid, and the other for protein. The particles are analyzed as they flow through a laser beam
hemagglutination assay
if antibodies are present, they stick to RBCs and cause mat formation. if antibodies are not present, the RBCs settle to the bottle forming a button
HA titer
the inverse of the greatest dilution that completely agglutinates the RBCs
high performance liquid chromatrophy (HPLC)
the concentrations of specific viral antigens may be quantified through UV analysis of fractions generated during this test. Gives indirect idea of virus concentration
single radial immunodiffusion (SRID)
radial diffusion of purified viral antigens (standards) and viral particles through agarose gel seeded with polyclonal antisera against a viral antigen
plaque
a circular zone of necrotic cells surrounded by viable cells in a biological assay
monolayer plaque assay
it is a functional measurement, and has no relation to the actual number of viruses. Units= PFU
virus replication cycle (10 steps)
attachment, penetration, uncoating, translation of early mRNA, translation of early proteins, replication of viral DNA, transcription of late mRNA, translation of late proteins, assembly of virions, release
determining titer (equation)
average plaque count x reciprocal of dilution selected
pock
necrotic region. Unit= pock forming units
transformation assay
quantitative determination of titers of oncogenic viruses. transformed cells lose contact inhibition and become heaped upon one another
quantal assay
measures presence of absence of infection
endpoint
virus dilution that affects 50% of the test subjects
tissue culture infectivity dose 50
dose in which 50% of the tissue dies
lethal dose 50
the dose that kills 50% of the subjects
embryo lethal dose 50
dose in which 50% of the embryos die
paralytic dose 50
dose that causes paralysis in 50% of the subjects
TCID50
the tissue culture infectious dose which will infect 50% of the cell monolayers challenged with the defined inoculum
multiplicity of infection (MOI)
the average number of virus particles infecting each cell
equation for TCID50
[(%infected above 50%) - (50%)] / [(%infected above 50%) - (%infected below 50%)]
