Dx tests (based off chart)

Question 1

Dx by gross evaluation and histopathology

Answer 1

Fixation with 10% neutral buffered formalin
Consult with lab tech
Case history + description on lesions

Question 2

Inclusion bodies

Answer 2

Nuclear or cytoplasmic aggregates of stainable proteins
Sites of viral multiplication (capsid proteins)
Examined with light or EM

Question 3

Which tests are used for Abs

Answer 3

ELISA, WB, HAI, immunodiffusion, neutralization, IgM specific

Question 4

T/F: Tissue fixation or viruses bound to Abs can mask detection and lower sensitivity of the assay for IF detection

Answer 4

TRUE

Question 5

ELISA

Answer 5

Tissues/ serum
Fast
High sensitivity, low specificity (false +)

Question 6

When do you use ELISA?

Answer 6

Target large # of sick animals
Targets Ab and Ag

Question 7

Advantages of ELISA

Answer 7

Rapid (few hours)
15 mins for a single animal
Automated equipment for many

Question 8

Disadvantages of ELISA

Answer 8

Specificity towards altered viral ags

Question 9

Commercial ELISA kit (SNAP)

Answer 9

Detects FeLV (confirmed by IFA) and FIV Ab in blood (confirmed by WB with unvx animals)

Question 10

What are serological tests used for?

Answer 10

Detect prior infection by a virus
Detect immune response to vx
Detection of response to viruses in acute (blood IgM titer) and chronic phases (IgG)
Pair sampling
Distinguishing Abs response as a result of natural infection v. vx

Question 11

Western Blot (immunoblotting)

Answer 11

Serum
Slow and qualitative
Transfer of proteins from gel surface to a special membrane that capture proteins

Question 12

When do you use WB?

Answer 12

Confirmation of Ab test (FIV)

Question 13

Hemaggluntination test (HAI)

Answer 13

Serum
Rapid
Sensitive and specific

Question 14

When is HAI used?

Answer 14

Any virus that agglutinates RBCs ( influenza or parainfluenza)

Question 15

Immunodiffusion

Answer 15

Serum and tissue samples (oldest)
Slow (24-48 hrs)
Specific but not sensitive, use with ELISA

Question 16

Advantages of immunodiffusion

Answer 16

Simple, inexpensive, doesn’t require production of infectious virus

Question 17

Disadvantages of immunodiffusion

Answer 17

Qualitative assay (yes/no), lacks sensitivity

Question 18

Neutralization

Answer 18

Serum
Slow and expensive
Detects and quantifies virus specific Abs in blood

Question 19

When is neutralization used?

Answer 19

Testing efficacy of vx titer
If pet was recently sick
Prevent virus infection of cells by Abs present in blood

Question 20

What is the most common procedure of virus neutralization test

Answer 20

Constant virus variable serum method

Question 21

Advantage of virus neutralization test

Answer 21

Species independent

Question 22

Disadvantages of virus neutralization test

Answer 22

Slow: takes a few weeks to grow a virus
Labor intensive: growing and maintaining cell culture

Question 23

IgM specific

Answer 23

Serum
Rapid
Sensitive and specific

Question 24

When is IgM specific used?

Answer 24

IgM: acute, less cross reactive with other viruses
IgG: chronic

Question 25

Tests for Ags

Answer 25

ImmunoEM (IEM), Immunofluorescent assay (IFA), ELISA and immunoperoxidase

Question 26

ImmunoEM (IEM)

Answer 26

Samples from where virus is shed Tissues, cells, secretions and excretions Fast but expensive More sensitive than EM

Question 27

When is IEM used?

Answer 27

Dual infections Low virus concentrations Non-cultivated viruses

Question 28

Immunofluorescent assay (IFA)

Answer 28

Frozen fixed tissue and liquid nitrogen Fast (3 hrs) High sensitivity (more specificity than ELISA)

Question 29

Disadvantages of IFA

Answer 29

Requires special microscope and cryostat for tissue sectioning of frozen samples Can't ID all viruses (enterovirus)

Question 30

Immunoperoxidase (IP)

Answer 30

Frozen tissue, has longer shelf-life Slow >25 hrs Sensitive and specific

Question 31

Advantages of IP

Answer 31

Examined under LM Duration and stability of signal Better cellular localization of virus in tissue

Question 32

When is IP used?

Answer 32

Better localization of virus (intracytoplasmic or intranuclear)

Question 33

PCR (nucleotide)

Answer 33

Fast (2-8 hrs) High sensitivity and high specificity

Question 34

When is PCR used?

Answer 34

Viruses can't be cultured readily Non-infectious viruses Undetectable viruses Viruses complexed to Abs Enteroviruses

Question 35

Cytopathic effects

Answer 35

Swelling, rounding, shrinking, syncytium formation, clustering, complete destruction of the monolayer

Question 36

Cell cultures

Answer 36

When other tests aren't possible (new pathogen) Vx manufacturer If viruses causes disease in a sample of several viruses Required to examine sublet CPE Requires confirmatory testing

Question 37

_________ is the most accurate of the quantativative biological assays

Answer 37

Plaque assay

Question 38

Plaque assays

Answer 38

Cytopathic viruses produce holes in cells monolayer → each virus infectious particle produces a circular visible eye clear zone (plaque= dead cells)