Diagnostics introduction Flashcards
Molecular diagnostics is a qualitative or quantitative measure of what 2 things?
- DNA or RNA
- Proteins
What does the choice of diagnostic test depend on?
(5 things)
- the type of sample
- clinical question (are you screening or diagnosing – antibody vs antigen)
- availability of test
- intended use
- performance characteristic of test (sensitivity, specificity, PPV, NPV, reliability)
What are 4 DIRECT diagnostic testing methods?
- electron microscopy
- genome identification (PCR)
- antigen detection
- bacterial culture
Why is virus isolation considered an INDIRECT testing method as opposed to a direct method?
virus isolation is looking at the EFFECT of the virus on the cell culture.
what type of diagnostic method is used for antibody detection IgM and IgG?
serology
What 7 tests allow for the detection of ORGANISMS?
- light microscopy
- electron microscopy
- ELISA
- IFA
- IHC
- PCR
- NGS
What diagnostic test is good for detecting organisms that cause diarrhea, such as coronavirus or rotaviruses, even in non-clinical animals.
electron microscopy
What is the biggest downside to virus isolation and is the reason it is not readily performed unless in a state lab with lots of resources?
virus isolation requires cell cultures which is expensive and takes lots of time.
What 3 diagnostic tests detect antigen (protein)?
- ELISA
- Immunofluorescence (IFA)
- Immunohistochemistry (IHA)
What is the key component in ELISA that allows for the color change to occur?
substrate is added and the enzyme converts it into a detectable product. a color change occurs during this process.
____________ uses cell culture or tissue samples. Antibodies have fluorescent tags, so they fluoresce under microscopic light.
immunofluorescent antibody test (IFA)
T/F: IFA tests are extremely sensitive and can detect low viral titers.
false – they require high viral titers.
__________ uses paraffin-embedded tissues mounted on glass slides. The samples are incubated with specific antibodies tagged with a substrate (chromogen) to give a color reaction. This test allows for the detection of antigen IN THE LESION or in specific cells.
immunohistochemistry
What is the most commonly used diagnostic test to diagnose neoplasia?
Immunohistochemistry
Immunohistochemistry is characterized by ________ against tumor markers to detect: cells of origin, stage of differentiation, and prognostic indicators.
antibodies
What 3 diagnostic methods are used for genomic detection?
- PCR
- in-situ hybridizaton
- next generation sequencing
PCR requires _______ and probes that are specific to the target DNA. PCR also requires that you have ENOUGH, GOOD QUALITY target DNA/RNA to amplify.
DNA primers
If you get a negative canine parvovirus PCR test, but you are sure that this patient is positive for parvo, what could have occured?
the virus was not detected by the PCR test because the non-conserved DNA portion was picked up.
When there is a LOT of DNA/RNA present, what can you expect about the signal on a PCR test?
brighter fluorescence
How are we able to use PCR for neoplasia?
PCR can identify clonal lymphocyte populations (PARR), detect chromosomal translocations, deletions, or duplications, and can detect c-kit mutations in mast-cell tumors
If you have a sample that has lots of small lymphocytes and you are trying to differentiate between neoplasia (lymphoma) or inflammation, how could you make this diagnosis?
with neoplasia, all of the lymphocytes will have the same receptors. In inflammation, the cells will have variable receptor types.
__________ allows for visualization of virus in tissue or at the site of infection. It can help us to determine which virus is responsible for pathologic lesions.
In-situ hybridization
What diagnostic test can help determine unknown viruses?
next generation sequencing
Why can next generation sequencing be tricky?
It gives you back a bunch of different organism information. It is us to you as the doctor to take the information and combine it with the patients clinical signs to determine which pathogen is causing the disease present.
What diagnostic tests are good for detecting antibodies?
Serology – ELISA, immunodiffusion, and virus neutralization
What is the purpose of ELISA dilutions?
detect antibody titers
Why is it important detection of IgM important?
tells you that this patient is in early stages of disease.
__________ is characterized by antigen being loaded into the center of the gel and test serum being added onto the periphery. A charge gets added and antibodies (if present in your sample) then move toward the antigen and precipitate creating a band.
immunodiffusion
Virus neutralization is characterized by using ________ in serum to inhibit virus replication
antibodies
A negative cell culture with virus neutralization indicates what?
+ for whatever you’re looking for.
ie if you are testing for equine herpes virus, and your virus isolation test has no cell growth, then this patient is positive for equine herpes virus.
T/F: antibodies will decrease without continouus exposure.
true
what are possibilities for why a test may not give you the expected results ?
- technical problem
- early infection or in recovery
- false + (low prev)
- sequestered in tissues that were not tested.
_________ is best used when determining what organisms are circulating in a population.
serology
