Dx tests (based off chart) Flashcards
Dx by gross evaluation and histopathology
Fixation with 10% neutral buffered formalin
Consult with lab tech
Case history + description on lesions
Inclusion bodies
Nuclear or cytoplasmic aggregates of stainable proteins
Sites of viral multiplication (capsid proteins)
Examined with light or EM
Which tests are used for Abs
ELISA, WB, HAI, immunodiffusion, neutralization, IgM specific
T/F: Tissue fixation or viruses bound to Abs can mask detection and lower sensitivity of the assay for IF detection
TRUE
ELISA
Tissues/ serum
Fast
High sensitivity, low specificity (false +)
When do you use ELISA?
Target large # of sick animals
Targets Ab and Ag
Advantages of ELISA
Rapid (few hours)
15 mins for a single animal
Automated equipment for many
Disadvantages of ELISA
Specificity towards altered viral ags
Commercial ELISA kit (SNAP)
Detects FeLV (confirmed by IFA) and FIV Ab in blood (confirmed by WB with unvx animals)
What are serological tests used for?
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
Western Blot (immunoblotting)
Serum
Slow and qualitative
Transfer of proteins from gel surface to a special membrane that capture proteins
When do you use WB?
Confirmation of Ab test (FIV)
Hemaggluntination test (HAI)
Serum
Rapid
Sensitive and specific
When is HAI used?
Any virus that agglutinates RBCs ( influenza or parainfluenza)
Immunodiffusion
Serum and tissue samples (oldest)
Slow (24-48 hrs)
Specific but not sensitive, use with ELISA
Advantages of immunodiffusion
Simple, inexpensive, doesn’t require production of infectious virus
Disadvantages of immunodiffusion
Qualitative assay (yes/no), lacks sensitivity
Neutralization
Serum
Slow and expensive
Detects and quantifies virus specific Abs in blood
When is neutralization used?
Testing efficacy of vx titer
If pet was recently sick
Prevent virus infection of cells by Abs present in blood
What is the most common procedure of virus neutralization test
Constant virus variable serum method
Advantage of virus neutralization test
Species independent
Disadvantages of virus neutralization test
Slow: takes a few weeks to grow a virus
Labor intensive: growing and maintaining cell culture
IgM specific
Serum
Rapid
Sensitive and specific
When is IgM specific used?
IgM: acute, less cross reactive with other viruses
IgG: chronic
Tests for Ags
ImmunoEM (IEM), Immunofluorescent assay (IFA), ELISA and immunoperoxidase
ImmunoEM (IEM)
Samples from where virus is shed
Tissues, cells, secretions and excretions
Fast but expensive
More sensitive than EM
When is IEM used?
Dual infections
Low virus concentrations
Non-cultivated viruses
Immunofluorescent assay (IFA)
Frozen fixed tissue and liquid nitrogen
Fast (3 hrs)
High sensitivity (more specificity than ELISA)
Disadvantages of IFA
Requires special microscope and cryostat for tissue sectioning of frozen samples
Can’t ID all viruses (enterovirus)
Immunoperoxidase (IP)
Frozen tissue, has longer shelf-life
Slow >25 hrs
Sensitive and specific
Advantages of IP
Examined under LM
Duration and stability of signal
Better cellular localization of virus in tissue
When is IP used?
Better localization of virus (intracytoplasmic or intranuclear)
PCR (nucleotide)
Fast (2-8 hrs)
High sensitivity and high specificity
When is PCR used?
Viruses can’t be cultured readily
Non-infectious viruses
Undetectable viruses
Viruses complexed to Abs
Enteroviruses
Cytopathic effects
Swelling, rounding, shrinking, syncytium formation, clustering, complete destruction of the monolayer
Cell cultures
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
_________ is the most accurate of the quantativative biological assays
Plaque assay
Plaque assays
Cytopathic viruses produce holes in cells monolayer → each virus infectious particle produces a circular visible eye clear zone (plaque= dead cells)