working conference 1

  1. What is ELISA used to detect?
    presence of Ab in pt's sera
  2. What are the steps for ELISA?
    • 1. Ag coated onto surface of plastic well
    • 2. pt's serum (w/ Ab) added to well; binding of Ab/Ag and excess washed away
    • 3. enz-labeled anti-Ab added to well to bind to pt's Ab; excess washed away
    • 4. colorless subst added to well; reacts w/ enz still there to produce a color
  3. What does the Western Blot begin w/?
    electrophoresis: separates mix of prot based on diff rates of migration in electric field (SDS-PAGE
  4. What happens after electrophoresis w/ Western Blot?
    pt antiserum incubated w/ blot, any Ab present that recog electrophoresed prot on blot bind; bound Ab can be detected w/ labeled anti-Ab to allow color rxns
  5. What is the basic principle of Western Blot?
    sep a standardized prepartion of Ag derived from a particular pathogen and demonstrate "pattern" of Ab reactivity that provides a fingerprint unique to that pathogen
  6. What are the adv of Western Blot?
    simultaneously demonstrate reactivities of mult Ab in polyclonal pt antiserum against mult Ag assoc w/ particular path
  7. Direct Fluorescenct Ab test
    primary Ab (of known Ag specificity and labeled w/ fluorescein) reacts w/ specific Ag in test specimen
  8. Indirect Fluorescent Ab Test
    presence of an unlabeled "primary" Ab bound to Ag in a clinical specimen is detected by fluorescein-labeled "secondary" Ab; secondary finds and binds to primary
  9. What is the adv of IFA?
    increased sensitivity of detection due to amplification effect as mult secondary Ab can bind to each single molecule of primary Ab
  10. precipitation rxn
    dep on bivalent characteristics of Ab that allows it to crosslink mol of Ag to each other; sol Ab bind to sol Ag molecules and lead to precip of lg insoluble Ag:Ab complexes...agglutination
  11. RPR card test
    agglutination test to detect reagin-type reactivity; uses carbon particles that are coated w/ cardiolipin Ag that reacts w/ reagin
  12. reagin
    Ab that react w/ lipid Ag assoc w/ Treponema
  13. tube agglutination
    determines serum Ab titers in diagnosis of infectious diseases; demonstrates agglutination in tubes and prozone effect
  14. prozone effect
    conc of Ab is too high in relation to Ag (Ab interferes w/ its ability to crosslink); serum that is agglutination neg bc of prozone becomes pos when diluted
  15. complement fixation test
    Ag:Ab rxn can lead to fixation of complement which triggers complement cascade and leads to cell lysis
  16. What is a positive CF test?
    complement binds to Ag:Ab complex; no lysis of RBC bc no free complement in serum bc all bound to Ag:Ab complex
  17. What is a negative CF test?
    no Ab present so no Ag:Ab complexes; free complement binds to RBC antibody and MAC forms
  18. Neutralization (Virus Plaque Inhibition) Assay
    monolayers on culture plates exposed to pt specimens that may have virus; if virus in pt serum then it will infect cells in monolayer and make them lyse (CPE)...plaques form
  19. no CPE
    viruses cannot infect monolayer bc of presence of virus-specfic Ab; no plaques; neutralizing Ab indicative of an Ab response against particular virus
Card Set
working conference 1
MS1/Mod 1: Immunology; serologic diagnosis