Virus Quantification

Question 1

What is virus quantification?

Answer 1

counts the number of viruses in a specific volume to determine the virus concentration

Question 2

What is the virus titer?

Answer 2

lowest concentration of a virus that still infects cells (# infectious units per mL sample)

Question 3

What are the 2 types of viral quantification tests?

Answer 3

• biological: depends on a virus particle initiating a successful replication cycle
• physical: don’t depend on any biological activity of the virus particle

Question 4

When is direct particle counts by TEM ideal?

Answer 4

for viruses with unique geometrical shapes

*expensive, requires training, need standard particle of known concentration

Question 5

What is the Virus Counter 2100 based on?

Answer 5

flow cytometry for use with nanometer scale particles

*can measure intact virions through detection of co-localized protein and nucleic acids

Question 6

What are the two stains used for the Virus Counter 2100?

Answer 6

one for nucleic acid, one for protein -> then analyzed as they flow through laser beam

Question 7

HA titer

Answer 7

inverse of greatest dilution that completely agglutinates the RBC

Question 8

T/F if there is no hemagglutination, RBC will settle as a button

Answer 8

T: if there is hemagglutination RBC will form a mat

Question 9

High Performance Liquid Chromatography (HPLC)

Answer 9

concentrations of specific viral antigens may be quantified through UV analysis of fractions generated during HPLC

Question 10

single radial immunodiffusion (SRID)

Answer 10

radial diffusion of purified viral antigens (standards) and viral particles through agarose gel seeded with polyclonal antisera agaisnt a viral antigen

• antigen starts diffusing from center out
• point where antigen conc = antibody conc you will see a circle

Question 11

T/F with SRID, the larger the diameter of the circle, the lower the concentration of virus (and antigen)

Answer 11

F: larger circle = higher conc of virus

Question 12

qPCR

Answer 12

utilizes PCR chemistry to amplify viral DNA or RNA to produce high enough concentrations for detection and quantification by fluorescence

Question 13

What is the most accurate of the quantitative biological assays?

Answer 13

monolayer plaque assay

Question 14

PFU

Answer 14

measures the number of virus particles capable of forming plaques per unit volume

Question 15

plaque

Answer 15

no relation to actual number of viruses; few cells that have been destroyed by virus before agar restricts it

Question 16

pock

Answer 16

necrotic area on chorioallantoic membrane (CAM) of embryonated egg *titration of herpesviruses and poxviruses

Question 17

transformation assay

Answer 17

quantitative determination of titers of oncogenic viruses

*units: focus-forming units/mL

Question 18

Do transformed cells have contact inhibition?

Answer 18

No, they become heaped upon each other

Question 19

quantal assay

Answer 19

measures presence of absences of infection; used for certain viruses that do not form plaques or for determining virulence of a virus in animals or eggs

Question 20

endpoint

Answer 20

virus dilution that affects 50% of test subjects

Question 21

lethal dose 50

Answer 21

dose that will kill 50% of subjects

Question 22

TCID50

Answer 22

tissue culture infectious dose which will infect 50% of cell monolayers challenged with inoculum

Question 23

multiplicity of infection (moi)

Answer 23

average number of virus particles infecting each cell

Question 24

Reed and Muench method

Answer 24

TCID50 = (%infection above 50% - 50%) / (% infection above 50% - %infection below 50%)

*add this proportional factor to dilution next above 50%