Study Notes on Enzymes and Immunofluorescence Assays

Immunofluorescence Assay: Uses fluorescent molecules (fluorophores) instead of enzymes to detect antigens.
Cost Efficiency: More economical than radioactivity, less so than ELISA.

Direct Immunofluorescence:
- Detects antigens directly in patient tissue samples (e.g., biopsy).
- Antibody conjugated with a fluorescent tag binds directly to the antigen.
- Requires fluorescent microscopy for detection.
Indirect Immunofluorescence:
- Involves a wash step and uses patient serum over reagent cells.
- Secondary fluorescent antibody binds to captured human antibodies.
- It is used to identify autoimmune antibodies (e.g., ANAs).

Competitive Assay: Results inversely proportional to antigen presence.
Polarized light emitted when bound to an antibody; more antigen means less fluorescent labeling.
Advantages: Simple implementation, minimal hazardous waste.
Disadvantages:
- Susceptible to signal interference from patient serum substances.
- Requires specialized equipment for polarization.

Emission of light from enzyme reaction with substrates (e.g., luminol and hydrogen peroxide).
Advantages:
- Low cost and long shelf life.
- Fast detection speed, sensitive, comparable to RIA and EIA.
Disadvantages:
- Precision in injectors needed for consistency.
- Potential for false positives due to quenching from urine/plasma.
- Requires dedicated light detection analyzer for results interpretation.