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22 spring: immunology > Test 1: lecture 12 and 13 testing > Flashcards

Test 1: lecture 12 and 13 testing Flashcards

1
Q

antibody test shows evidence of ___

A

PIE (protection (vaccine), infection or exposure)

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2
Q

two main ways to make definitive diagnosis of an infection with an antibody test

A

4-fold increase in titer of non-class specific Ab

or

Detection of IgM antibodies (ideally paired with IgG)

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3
Q

___ is the highest dilution of an assay to give a positive reaction

A

titer

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4
Q

•Detecting a four-fold rise in titer requires___

A

acute and convalescent titers

2 weeks apart

(titer is the highest dilution that gives a positive reaction→ at lower dilutions they clump together and fall to bottom, at higher dilutions there are so little that they are all spread out, not clumping)

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5
Q

are titers absolute or relative value

A

relative (has no unit)

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6
Q

elevated liver and kidney values are common with ___

A

lepto

run agglutination antibody titer assay and again 2 weeks later

4x increase is positive for lepto

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7
Q

___ indicated recent exposure or infection

A

IgM

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8
Q

___ represents previous exposure

A

IgG

(secondary response)

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9
Q

antigen test

A
  • Detects a SPECIFIC antigen of a pathogen.
  • In the presence of clinical disease detection of an antigen is considered diagnostic.
  • Can cross react with similar targets (from closely related pathogens for example).

antigen titers can increase with treatment

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10
Q

what is one down fall of the antigen test

A

pathogens can be very closely related

therefore antigen test can have cross react → react to wrong antigen

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11
Q

weird shape of nose, run cryptococcus ___ test and it is + at 1:8. Treat. Rerun test and it is + 1:16. why the increase after treatment?

A

antigen test

by treating the fungus, it is broken down leading to more antigen and more response by body = swelling and higher antigen test results

antigen titers can rise with treatment

treat until you get 0 on titer

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12
Q

what does a negative antigen test show

A

pathogen not there

antigen present, but not enough to detect

antigen present but bonded to antibody

technical error

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13
Q

what does a + antigen test show

A

pathogen present

antigen present but viable microbe absent

cross reactive antigen present from another pathogen

technical error

poor assay specificity (false +)

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14
Q

negative antibody test →

A

exposure to pathogen did not occur

too early in course of infection

severe immunosuppression

technical error

poor assay sensitivity (false negative)

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15
Q

positive antibody test _>

A

previous natural exposure to pathogen

previous vaccination

cross- reactive

technical error

poor assay specificity → false +

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16
Q

seronegative

A

used to screen blood donor

immunoassay studies are 0

antibody test → dog was never exposed to this pathogen and carries no antibodies for these diseases

17
Q

what kind of test do you use for blood donors and why?

A

antibody→ cast broad net

will show if every exposed, previous immunization

don’t use antigen→ cause antigens at very low levels can’t be detected and some diseases can lead to long term effects that would exclude the patient from being a blood donor

18
Q

why use titer for vaccination status

A
  • Assess status prior to booster
  • For Animals with Concurrent disease
  • Assess immunity at > 24 weeks
  • Unknown vax hx
  • Assess prior to breeding
  • To determine if animals are genetic “non-responders”
  • Test in face of outbreak for quarantine purposes
19
Q

IFA (immunoflourescent Ab)

A

slide with antigen are covered with antibodies that have been linked to specific fluorescent, if the smear contains the antigen the corresponding antibody will bind, wash away anything unbound, use dark field microscopy to see what sticks

  • Used to detect presence of a specific antigen directly in a sample (tissue or smear).
  • Slide is incubated with fluorescently labeled Ab and the unbound Ab washed away.
  • Darkfield microscopy with ultraviolet light is use to detect bound antibody.
  • Can also be used for tumors, cell types etc.
20
Q

___ is a test where a slide with antigen are covered with antibodies that have been linked to specific fluorescent, if the smear contains the antigen the corresponding antibody will bind, wash away anything unbound, use dark field microscopy to see what sticks

A

IFA (immunofluorescent antibody)

21
Q

ELISA

A

enzyme linked immunosorbent assay

•Either Bidirectional Flow ELISA or Immunochromatography for in-house assay.

22
Q

indirect elisa

A

antigen bound to bottom of well

add serum from patient (may contain antibody)

wash away extra stuff

enzyme linked secondary antibody (or antiglobulin→ antibody that binds to antibody)

wash away

add chromogenic substrate that binds to secondary antibody

color change→ antibody was in serum = +

23
Q

sandwich ELISA

A

•Capture Ab is coated on plate and Ag in sample is bound.••Unbound antigen is washed a detection Ab is added.••Finally a labeled antiglobulin is added for detection.

sandwich looks for antigen

capture antibody is coated on bottom of plate

add serum that might have the antigen in it

wash away every thing else

add enzyme linked antibody to that antigen we are looking for

wash everything else away

add labeled anti-globulin ( antiglobulin→ antibody that binds to antibody) that reacts to 2nd antibody(changes color)

24
Q

what type of ELISA is for antigen identification

A

sandwich

•Capture Ab is coated on plate and Ag in sample is bound.••Unbound antigen is washed a detection Ab is added.•Finally a labeled antiglobulin is added for detection.

25
Q

•Capture Ab is coated on plate and Ag in sample is bound.••Unbound antigen is washed a detection Ab is added.••Finally a labeled antiglobulin is added for detection.

A

sandwich elisa

looking for a specific antigen

26
Q
  • Ab is detected in sample when it is bound to antigen that is coated on the plate.
  • Plates are washed of unbound Ab.
  • Secondary Ab (or antiglobulin) with reporter is bound to sample Ab.
A

indirect ELISA

(looking for antibodies)

27
Q

Bidirectional flow ELISA

A

snap test

matrix is pre-coated with specific antibodies

add sample, that has enzyme linked antibodies added to it

if antigen is present in sample will bind to the antibodies on the matrix and the enzyme linked antibodies will keep it from getting swept away

the substrate then moved across the cleaned matrix and blue dot = +

28
Q

competitive ELISA

A

color change is inversely proportional to AB or Ag level

no color change = +

well is bound with antigen, add sample, if sample has antibodies will bind to the bound antigen, then add lab derived enzyme linked antibodies,

if there are no antibodies in the original sample lab Ab will bind and if there was antibodies in the original sample lab Ab will not bind

if lab sample Ab bind there will be a color change = negative

29
Q

___ has color change is inversely proportional to Ab level in serum

A

competitive elisa

negative result = color change

positive result = no color change

30
Q

why use competitive elisa

A

not species specific

other antibody ELISA need antiglobulins that are against the species Ig

where competitive just needs lab derived antibody

31
Q

AGID

A
  • Based on the passive diffusion of soluble Ab and Ag through agar gel.
  • When “positive” precipitation occurs.
  • Inexpensive but subjective.

4= negative

put lab derived antigen in the center, in wells around put +/- control and serum

precipitation line = +

32
Q

serum neutralization

A
  • Typically used for viral diagnosis.
  • Observation of virus challenged cell cultures in the presence of test serum.
  • If cells do not become infected then there is antibody in the serum.
  • Results can be titered.

have specific cell line, add in virus, if the cells become infected → no antibodies in cell line. If cells do not become infected → + antibodies

33
Q

have specific cell line, add in virus, if the cells become infected → no antibodies in cell line. If cells do not become infected → + antibodies

A

serum neutralization

used for viral diagnosis

34
Q

complement fixation

A

if serum has antibodies, RBC will pellet and no lysis

have serum, heat it up to kill complement

add lab derived antigen, if serum has antibodies already present will bind

then will add lab derived complement

if Ab:Ag present, complement will bind

then add in sheep RBC and antibodies to sheep RBC

if complement is bound because antibodies are present in original sample→ RBC will settle into a pellet, no lysis (complement was used up → nothing happens to added RBC)

35
Q

have serum, heat it up to kill complement

add lab derived antigen, if serum has antibodies already present will bind

then will add lab derived complement

if Ab:Ag present, complement will bind

then add in sheep RBC and antibodies to sheep RBC

if complement is bound because antibodies are present in original sample→ RBC will settle into a pellet, no lysis (complement was used up → nothing happens to added RBC)

A

complement fixation test

22 spring: immunology (23 decks)

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