(Exam 2) MLS 420 Immunoassay Methods part 2

immunofixation electrophoresis (IFE)

This is a precipitation method that uses the detection of monoclonal antibodies or immunoglobulins in the production of one antibody isotope, multiple myeloma, or Waldenstroms.

CSF, urine, serum

What are the most common specimens used for immunofixation electrophoresis?

gamma region

IFE is a followup test t serum protein electrophoresis to understand why there is an increase in the ______ of SPE.

separated, antibodies, precipitate

For IFE methodology: the proteins are ______ by electrophoresis, then know anti-human _____ applied to the gel, and the immune complexes form that _____ out of the gel.

antigen

For IFE, the human antibody acts as the ______.

agrose gel

The two stages of process using ______ in IFE are electrophoresis and immunoprecipitation.

IFE

The Western Blot method is in the ______.

Western blot

This test can produce qualitative or semi-quantitative information about proteins and uses antibodies directed against the protein.

blotting

______ is the transfer of biological material from a gel onto a membrane.

confirmatory

Western blot is a _____ test for HIV1, Lyme's Disease, Hep B, Herpes, etc...

size

The Western blot is a separation of proteins based on _____ by electrophoresis method.

membrane, patient

The antigen in the _____ and the _____ antibody forms the immune complex for Western blot.

fluorescent immunoassay

This is a labeled immunoassay that has the fluorescent tag that absorbs energy from incident light that must be converted to a light of longer wavelength and light emitted is then measured.

fluorescein (green), tetramethyrhodamine (red)

For the Fluorescent immunoassay tags, _____ aborbs at 490 and emits at 520, and the _______ aborsbs at 590 and emits at 580.

low wavelength, longer wavelength

Fluorescent immunoassays absorb energy from a ______ of light and then a photon of ______ of light is then released.

direct, indirect, and fluorescent polarization

What are the three methods of Fluorescent immunoassays?

direct immunofluorescence

This method of fluorescent immunoassays is looking for the antigen in the patient sample using a single reagent labeled antibody directed to the target of interest in the patient sample.

heterogenous solid phase

Direct immunoflorescene is a _______ separation method that the specimen is fixed to a slide and is then washed to remove any unbound reactants.

specific and sensitive, cross-reactivity

The direct immunofluorescence test has the advantages of: simple, quick, _______, low cost. The disadvantage is: ______, subjectivity of reading the slide, and lower signals.

bacterial/viral

The direct immunofluorescence assay is used for the detection of _______ antigens like legionella and tissue antigens from biopsies.

indirect immunofluorescence

This florescent immunoassay is looking for antibodies in the patient sample using a known florescently labeled reagent antibody directed against the analyte of interest in the patient.

unlabeled, heterogenous

The indirect immunoflouroescence uses a known _____ antigen attached to a solid phase ______ method.

initial immune complex

The indirect immunofluorescence assay is indirect because the labeled reagent is not involved in the ______ formation.

twice

The indirect immunofluorescence assay is washed ______.

fluorescence

If the patient serum contains the antibodies in indirect immunoflorescnece assay, ______ will be detected.

indirect (secondary reagent amplifies)

What assay has greater sensitivity for immunofluorescence?

rubella

The indirect immunofloresence assay is used for bacterial or viral antibodies like syphilis and ____ and the antinuclear antibodies that cause autoimmune disorders.

fluorescent polarization immunoassay

This fluorescent immunoassay is homogenous, competitive assay with how fluorescent molecules behave in solution when excited by polarized light and measures how long it takes a fluorescent molecule to rotate.

emit light

In the fluorescent polarization immunoassay, when exposed to polarized light, molecules that are not rotating ______ back into a fixed plane.

unpolarized light

Molecules and small molecules that rotate easily emit _______ not in a fixed plane in the fluorescent polarization immunoassay.

polarized light

Labeled molecules that are bound to an antibody cannot rotate as fast, thus emitting more ______ in fluorescent polarizing immunoassay.

align

Incident light transmits light in all directions and once the light is transmitted through a polarized filter, the light waves _____ into one orientation either vertical or horizontal.

inversely proportional

The relationship for the fluorescent polarization immunoassay is ______ to the molecule volume and polarization.

sensitivity/specificity, auto-fluorescent

Advantages of the fluorescent polarization immunoassay has high ______ and versatile. Disadvantages are that they can _______ from other organic substances and are non-specific binding of serum substances.

therapeutic drug monitoring

The fluorescent polarization immunoassay is used for _____, drugs of abuse, hormones, and detection of food-borne toxins.

tag

For classifying the enzyme immunoassay, the assay on the _____ is the reagent. Therefore, if the tag is enzyme it is an EIA.

enzyme immunoassay

This is a labeled immunoassay with the enzyme tag attached to reagent antibody or antigen that measures the color, fluorescence, or luminsence produced by the enzyme acting on the substrate.

patient analyte

In the EIA, the labeled reagent reacts with unlabeled ______ if present to form the immune complexes.

substrate (not the enzyme)

What determines if the product produced is chromogenic, fluorescent, or luminescent?

sensitivity, speed, interfering

The choice of enzyme depends on _____, ease and _____ of detection, stability, availability, cost, absence of ______ substances.

catalyst

Enzymes are _____ that are not used up in the reaction and fact with a substrate to create a product.

substrate

This is the substance that the enzyme acts on/what the enzyme is biologically reactive with.

solid phase

Enzyme immunoassays are often performed in a _____ where reagent antigen or antibody is attached to micrometer wells, tubes, or beads.

conjugate antibody

This is an antibody that is linked to a label.

analyte

This is what is detected in the patient.

safe, temperature sensitive

The EIA advantages are: relative _____, cheap, sensitive, adaptable to automation. The disadvantages are: natural inhibitors, ______, non-specific binding, and enzyme label sites.

ELISA

This is the Enzyme linked immunosorbent assay that is the same methodology as EIA.

heterogenous competitive EIA

This is the _______ assay that has enzyme labeled reagent antigen competing with unlabeled patient antigen for binding sites on ta known reagent antibody that is attached to a solid surface.

inversely proportional

The heterogenous competitive EIA level of enzyme activity is ______ to the concentration of the patient analyte.

stop reagent

The ______ is the thing that stops any further enzyme action on the substrate.

heterogenous non-competitive EIA

This is the ______ immunoassay that has labeled reagent added in separate steps that is attached to a solid surface.

directly proportional

The heterogenous non-competitive enzyme activity is _____ to amount of analyte in the specimen.

heterogenous capture (sandwich) EIA

This is the ______ that is non-competitive with two reagent antibodies that capture antibody attached to a solid surface tagged with an enzyme that has two antibody outer layers with an antigen inside.

directly proportional

The heterogenous capture (sandwich) enzyme activity is ______ to amount of antigen.

antigen

In the heterogenous capture (sandwich), we are looking for the _____ in the patient.

heterogenous microparticle enzyme immunoassay

This is the ______ with a solution coated with known antibody with alkaline phosphatase labeled reagent antibody also added that measures fluorescence from the enzyme substrate interaction.

unknown antigen, non-competitive

The heterogenous microparticle enzyme immunoassay has the patient analyte as the _____, is automated, measures _____, fertility, cancer, metabolic, hepatitis, and thyroid markers and is __________.

directly proportional

The heterogenous microparticle enzyme immunoassay enzyme is ______ to analyte concentration.

homogenous competitive

This is the _______ with no separation steps, enzyme activity directly proportional to analyte level in the patient, detects hormones, drugs of abuse, and therapeutic drugs.

automate, sensitive

The advantages of homogenous competitive EIA is that it is easy to _____ and fast turnaround time. The disadvantage is that it is less ______ than heterogenous assays.

enzyme-multiplied immunoassay technique (EMIT)

This is a homogenous competitive assay technique that has known reagent antibody plus patient sample that is incubated to allow for immune complex formation then addition of a substrate.

enzyme

The EMIT in homogenous competitive assays measures _____ activity .

analyte (antigen)

Increased enzyme activity in the EMIT homogenous competitive assay means the ______ is present in the patient. and is directly proportional.

competes, stopped

In the EMIT competitive homogenous assay, the patient and tagged antigen _____ for binding sites but when the antigen binds to the free reagent antibody, the enzyme tag is _____ or inhibited from reacting with the substrate.

clinical toxicology

_____ is important with EMI, like therapeutic drugs, drugs of abuse, qualitative/quantitative on solid phase or solution.

chemiluminescent immunoassay (CIA)

These use chemical labels that, when oxidized, produce a substance of a higher energy level and when this substance decays it emits energy in the form of light.

heterogenous or homogenous

Chemiluminescent assays can be ______ and the tag attaches to the antibody or antigen to measure the light produced.

luminol

What is the best chemiluinescent that can be used?

sodum or hydrogen peroxide

What are the catalysts in CIAs?

fast turnaround time, tag

The advantages of CIAs are they have high sensitivity, inexpensive, _____ easy to use. The disadvantage is that the instrument is specific only to the _____, background signal interference and false results due to lack of precision.

rapid immunoassays

These are lateral flow immunoassays that are used in point of care testing and in lab settings with results within 30 minutes.

cartridge, dipstick

Rapid immunoassays can be in a _____ or _____.

nitrocellulose

What is the membrane for rapid immunoassays?

conjugate pad

The labeled reagent antigen or antibody is the _______ in the rapid immunoassays.

detection zone

The unlabeled reagent antigen or antibody is in the _____ in the rapid immunoassays.

control

The detection zone has built in _____ line and sample interpretation line in rapid immunoassays,

conjugate pad

The principle of rapid immunoassays is that the patient sample is added to one end of membrane strip, the sample then migrates through the _____ that contains the labeled antigen/antibody in the pad, and if the patient has the analyte then it forms an immune complex with labeled reagents.

basically any

What specimens can be used in rapid immunoassay tests?

easy, lower sensitivity/specificity

The advantages of rapid immunoassays are that they are rapid, low-cost, and _____ to perform. Disadvantages are they may have _______ and are only qualitative results.

multiplex immunoassay

This immunoassay detects multiple analyses in one test, can be labeled immunoassay with fluorescent or chemiluminescent labels.

microbead suspension or spatially separated solid phase

What are the two formats for multiplex immunoassays?

planar format (spatially separated solid phase)

This specific multiplex immunoassay uses two reagents that are capture reagent/ligands that is most commonly an antibody (unlabeled) and detector antibody (labeled).

solid surface

Each ligand that is captured is immobilized on a _____ and is mapped and known on a specific micrometer plate and is different to a different analyte of interest in the planar format.

microbead suspension

This type of multiplex immunoassay is the capture ligands which are usually antibodies are attached to a fluorescently activated microbes that contains the capture ligands directed to a different analyte of interest.

flow cytometry

Detection of the fluorescent signal in the microbes assay for multiplexes occurs via _____.

paired sera

Acute and convalescent samples are in _____.

2-4

Acute and convalescnt samples are collected _____ weeks apart.

increase in titer

To confirm a diagnosis, there needs to be an ______ between acute and convalescent titer.

4-fold change

For a significant change, there is a titer of a ______.

higher

An active infection will have a convalescent titer _____ than the acute titer.

acute infection happening

An example of acute/convalescent titers: If the acute titer is 1:2 (2) and the convalescent titer is 1:16 (16), then there is _______ because there is at least a four fold change in titer.

IgM

What immunoglobulin is in active infections that is produced by naive B cells and expect an increase in titer from acute to convalescent samples.

IgG, symptoms

______ only can be used but would need to see an increase in titer from acute to convalescent sample along with presence of _____ and the perfect situation would be to test both IgM and IgG.

IgG

What immunoglobulin is considered the immune antibody that has long term protection that is from mom until about 6 months of age?

IgM

What immunoglobulin is produced in the primary acute infection phase that has short term protection.

IgM

Which cannot cross the placenta and is made by the infant?

16

If the titer is 1:16 we report that as: