Immunoassays and Automated Procedures Flashcards
Which Immunoassay step requires washing steps?
Heterogeneous Immunoassays
Solid Phase
Competitive/Noncompetitive
Which Immunoassay step does NOT require a washing phase?
Homogeneous Immunoassays
Liquid Phase
Competitive
Faster/Easier
What are the different labels that are applied in immunoassays?
Enzyme Immunoassay
Chemiluminescence
Fluorescent Substances
What is Chemiluminescence?
Light emission produced during a chemical reaction
What are ELISA or EIA designed to do?
Detect antigens/antibodies by producing an enzyme triggered color change
What are the general approaches of EIA?
Nonisotopic labels (Detect total IgM/IgG)
Catalytic properties of enzymes (Detects and quantitates reactions)
What qualifications must be met for an enzyme to be used in immunoassay? (NOT SAUCE)
High stability
Extreme specificity
Not be present in antigen/antibody during testing
How do we quantify a Noncompetitive Enzyme Immunoassay?
Amount of color change is proportional to amount of antibody
How do we quantify a Competitive Enzyme Immunoassay?
Amount of color change is inversely proportional to amount of antibody
What are Capture Enzyme Immunoassays designed to detect?
Specific antibody type
IgM
Toxoplasma IgM
What are Chemiluminescent labels used to detect?
Proteins
Viruses
Oligonucleotides
Genomic nucleic acid sequences
Competitive or Sandwich
How do we quantify a Capture Enzyme Immunoassay?
Amount of light emitted is inversely proportional to amount of analyte (antigen)
What can luminol, acridinium esters, peroxyoxaltes, dioxetanes, and tris rutenium be grouped into?
Chemiluminescent labels
What has been used as visual tags for the presence of antibody?
Conjugated enzyme labels:
HRP
ALP
Avidin-Biotin
Which assay uses a conjugated antibody to detect antigen-antibody reactions at a microscope level?
Direct Immunofluorescent Assay (DFA)
Which assay is used to confirm the specificity of the FA technique?
Inhibition Immunofluorescent assay (IIA)
What is used in detecting Listeria Monocytogenes?
SQUID Technology
What test is measured photometrically and is assessed by the amount of cloudiness or turbidity?
Nephelometry
What is Nephelometry used to measure?
Complement components
Immune complexes
Presence of antibodies
What is Nephelometry based on?
The reaction between the protein being assayed and a specific antiserum
What is known as the relationship between antigen and the measurement of a constant antibody concentration?
Heidelberger Curve
What is Flow Cell Cytometry based on?
Reaction of dyes with the cellular component of interest
Stained cells pass through laser
What is the Polymerase Chain Reaction (PCR)?
In vitro method that amplifies low levels of DNA
What are the three cycles in PCR?
DNA Denaturation (Separation)
Primer Annealing (Recombination)
Extension of DNA (Synthetization)
What is a disadvantage of PCR?
Prone to producing false positives
Highly Sensitive (good thing)
What are three important applications of PCR?
DNA amplification
Target sequence Identification
Labeled antisense probe synthesis
What can PCR analysis lead to?
Detection of gene mutations
Coronary artery disease
What kind of charge does DNA have?
Net Negative charge
Migrates towards the anode (positive)
What are the advantages of molecular genetic testing?
Faster turnaround time
Smaller sample size
Increased specificity and sensitivity
What is considered the gold standard of molecular methods?
DNA sequencing
Displays exact nucleotide sequence
Why is the Sanger method important?
It is the most frequently used DNA sequencing method
What is a modified PCR technique that uses fluorescence resonance energy transfer to quantitate DNA sequences?
Real-Time PCR
What is a modified PCR technique that is used in detection and quantification of HIV and CMV?
Nucleic Acid Sequence Amplification
Which blot technique uses specimen DNA that are separated by electrophoresis?
Southern Blot
Sickle Cell Anemia is seen
Which blot technique uses mRNA from the specimen and is separated by electrophoresis?
Northern Blot
Which blot technique separates proteins electropheretically?
Western Blot
