Assays For Viruses

Question 1

How are viruses grown

Answer 1

Grown in tissue culture

Then infect cells and incubated within cells

The cells are then harvested if infected

Question 2

What does CPE mean which some viruses do to cells they infect

Answer 2

Cytopathic effects

Question 3

What is MOI

Answer 3

Multiplicity of infection

Question 4

What does multiplicity of infection measure

Answer 4

Number of infectious only particles which infect a cell

Question 5

What would an MOI of 10 mean

Answer 5

10 infectious particles found within a cell

Question 6

Why isn’t Multiplicity of infection same as particle per cell

Answer 6

Not all viruses have cytopathic effects

Question 7

What type of distribution has MOI have

Answer 7

Poisson

Question 8

What is a plaque assay

Answer 8

Way of quantifying viruses using serial dilutions and agar

Question 9

Why is agar important in plaque assays

Answer 9

Stops new viral progeny infecting cells

Question 10

What does PFU mean found on plaque assays in agar

Answer 10

Plaque forming units (from a single viral particle)

Question 11

Why wouldn’t all viruses show on the plaque assay

Answer 11

Not all have CPE cytopathic effects

Question 12

Does plaque asssays take a long time

Answer 12

Yes

Question 13

What is the difference in plaque assay and focus forming assays

Answer 13

Focus forming don’t use agar and are faster

Question 14

What is used to detect viruses in focus forming assays

Answer 14

Immunofluorescence which attach to viruses

Question 15

How are cells dna detected in focus forming assays

Answer 15

Via counter stain

Question 16

Why wouldn’t a FFU be the same as a PFU

Answer 16

Some particles are FFU (show up in immunofluorescence)

They don’t necessarily cause CPE which is what needs to show in PFU

Question 17

What does end point dilution assays use to test how many particles present

Answer 17

They use dilutions until no CPE is detected

Question 18

What is TCID50 which also uses end point dilution assays

Answer 18

It’s the conc of virus needed to kill 50% of the cells

Question 19

How many cells killed if there’s 100 TCID50

Answer 19

50/100

Question 20

What is TCID50 used for to counteract plaque issues

Answer 20

Allows the detection of non plaquing/CPE viruses

Question 21

What is an issue with TCID50

Answer 21

Takes a long time

Question 22

Which microscopes are used for protein assays for detection of viruses and what does it allow to find

Answer 22

Electron microscopy

Allows finding of the MOI (infectious particles in cell)

Question 23

What does haemogglutination measure

Answer 23

Number or viruses relatively (viable and non viable)

Via the clumping of RBC causing cross linking

Question 24

How can viruses non specifically mostly cause rbc clumping

Answer 24

Via their GP eg influenza HA

Question 25

What would need to be counted to get absolute number or viruses in haemogluttination

Answer 25

How many particles via EM

Question 26

Why doesn’t haemogluttination measure infectivity

Answer 26

Because even if cells are infectious they can be reactive and cause rbc cross linking

Question 27

Name the 3 types of Elisa

Answer 27

Direct

Indirect

Sandwich

Question 28

How does direct Elisa work

Answer 28

Sample added with possible antigen

The primary antibody with an enzyme attached would bind if antigen present

When substrate added it reacts and causes positive for the viral antigen

Question 29

How is indirect Elisa different

Answer 29

It uses a secondary antibody conjugate which binds to the antibody already present bound to antigen if the person is positive

Question 30

What do sandwich Elisa detect in sanple

Answer 30

CAPTURE Antibodies for the virus

Question 31

What is added to test capture antibody in sandwich

Answer 31

The antigen

Which then if present and attached will cause another antibody to bind

This antibody detected by a tertiary antibody with enzyme present and substrate added