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Immunology > Lecture 9/10: Methods/Vaccines > Flashcards

Lecture 9/10: Methods/Vaccines Flashcards

1
Q

ELISA

A

Enzyme Linked ImmunoabSorbent Assay; Ag/Ab quantification

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2
Q

Flow cytometry

A

IDs specific cell types w/ lasers and fluorescent Abs

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3
Q

Agglutination

A

Cell/molecule clumping when Ab is bound

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4
Q

Analyte

A

Molecule you want to measure

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5
Q

Polyclonal Abs

A
  • Mix of Abs from different B cell clones
  • Most IgG -> poly clonal response
  • Recognize different epitopes on same pathogen
  • Change over time for different Ags or increased affinity
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6
Q

Monoclonal Abs

A
  • Abs derived from 1 clone
  • No affinity maturation or change in antigen
  • Consistent response for diagnosis and therapy
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7
Q

Creating mAbs

A

Mice: mix spleen cells w/ cancer cells -> fused immortal hybridoma B cell clone
Humans: isolate desired clones, fuse w/ immortal myeloma

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8
Q

Agglutination test

A

E.g. Coombs test; positive or negative
- Detects Abs to RBC Rh factor, anti-Hu IgG binds mom’s anti-Rh IgG -> RBC agglutination

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9
Q

Types of ELISA

A

Direct: detect/quantify Abs
Indirect (sandwich): detect/quantify Ags

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10
Q

Direct ELISA

A

Ag bound on plate, detected w/ Ab

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11
Q

Indirect (sandwich) ELISA

A

Ag binds surface-coated primary anti-Ag Ab -> wash -> secondary anti-Ag Ab w/ tag gives signal

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12
Q

Visualization with tags vs probes

A

Tags: enyme tag adds colored substrate e.g. HRP; ELISA, IHC
Probes: fluorescent probes allow direct visualization of fluorescence e.g. FITC, Rhodamine; ELISA, immunofluor., flow

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13
Q

Serology

A

Measuring analytes in serum (soluble portion of CLOTTED blood post-centrifuge; plasma = anti-coag. blood post-centrifuge)

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14
Q

Titer

A

Last dilution above background

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15
Q

Concentration

A

Actual Ag amount

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16
Q

Absorbance

A

Dye concentration

17
Q

Lateral flow assay

A

e.g. at-home COVID test
- Conjugate pad w/ primary labeled Ab
- Test line w/ unlabeled other primary Ab
- Control line w/ secondary Ab
- Flow through generates lines

18
Q

Western Blot

A

Separate proteins by weight, transfer to membrane, Abs to visualize

19
Q

IHC/IF

A
  • Visualize cell/tissue morph. w/ light or fluorescence microscopy
    1. Prep slide
    2. Add primary mouse anti-Ag Ab
    3. Add labelled secondary anti-Ab HRP
    4. Add substrate for enzyme
    5. Color visual
20
Q

Types of immunization

A
  1. Passive
  2. Active
21
Q

Passive immunization

A

Providing preformed Abs to patient (faster, rabies, antivenom treatment)

22
Q

Active immunization

A

Using own immune system to make Abs T cells (vaccines)

23
Q

Types of vaccines

A
  1. Live attenuated
  2. Conjugate
  3. Toxoid
  4. Pathogen subunits
  5. Dead whole organisms
  6. mRNA
  7. DNA
24
Q

Live attenuated vaccine

A

MMR, Rotavirus, Varicella, Influ. nasal, oral polio, BCG
- Pros: long lasting immunity, similar or same immune response, diverse Abs generated
- Cons: can’t give to immunocomp.; pathogenic reversion

25
Q

Conjugate vaccine

A

Hemophilus influ. type b, pneumococcal conjugate
Protein conjugated to polysaccharide -> T dependent response

26
Q

Toxoid vaccine

A

Diptheria, tetanus
Vaccine w/ toxoids similar to pathogen toxins; immune response cross-reacts and neutralizes actual toxin

27
Q

Subunit vaccine

A

Vax w/ part of pathogen i.e. purified protein, recombinant protein, polysacch., peptides, etc.
- Pros: safe, easier quality control
- Cons: might not work or be as long lasting

28
Q

Multivalent vaccines

A

Subunit w/ combo of different subunits in 1 for different strains.
Subunit req. proper config. for protection (e.g. RSV failure)

29
Q

Killed whole organism vaccines

A

Pathogen inactivated w/ chemicals, heat, etc.
e.g. hepatitis, rabies, some flu
Can be very immunogenic, v. effective

30
Q

Combo vaccines

A

Multiple methods e.g. DTaP
Toxoid: diptheria, tetanus
Subunit: acellular proteins

31
Q

mRNA vaccines

A

Moderna, Pfizer
Encapsulated mRNA facilitating DC uptake -> protein expression -> presentation

32
Q

Adenoviral (DNA) vaccine

A

J&J
Disabled adenovirus delivers spike protein DNA; more stable vs mRNA but less immunogenic

33
Q

Adjuvants

A

Vaccine agents that enhance immune response via innate immunity; stim. DC cytokine secretion
- IL-12 -> Th1 diff. -> B support
- IL-6 -> B cell prolif.
- IFN-gamma -> stim. class switch, Ab response
- Upreg. CD28 express.

34
Q

Clinical trial phases

A
  1. Pre-clinical
  2. Phase I
  3. Phase II/III
35
Q

Pre-clinical phase

A

Experimental animals for safety/efficacy

36
Q

Phase I trials

A

Safety + dose escalation; <10 pts often w/ dose escalation, open label, no vulnerable popns

37
Q

Phase II/III

A

Test for efficacy; more subjects, randomized, controlled, double blind

38
Q

Challenge study

A

Challenge immunized pts w/ live pathogen; helpful w/ low disease incidence

Immunology (7 decks)

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