Indirect Antiglobulin testing

Question 1

What are other names for indirect antiglobulin testing?

Answer 1

AHG, coombs, enhanced ab detection

Question 2

How do you confirm negative results?

Answer 2

use a saline control

Question 3

What must you do for a forwards rxn that is all positive?

Answer 3

Check with saline and confirm positives

Question 4

Frequencies:
Antigen
D ag
C ag
c ag
E ag
e ag

Answer 4

D 85 lower white
C 70 lower black
c 80 lower asian
E 30
e 98

Question 5

What do you do to enhance weak rxn?

Answer 5

put in the fridge for IgM

Question 6

Describe the overview of Anti-D briefly
ig
stim

Answer 6

IgG immune stimulated, warm reactive (incomplete Ab)

Question 7

Describe the coombs test
detects?

Answer 7

detects weak/incomplete (IgG) ab using IgM to complete hemagglutination

indirect test

Question 8

Describe the coombs rxn agglutination test
what forms a lattice
must have removal of?
incubation?
method?

Answer 8

IgG ab forms a lattice w/ IgM Anti-IgG rgn
must have removal of unbound material
incubate at body Temp
INDIRECT

Question 9

Describe the immediate spin agglutination test
What forms a lattice
centrifuge
Plasma to cell ratio
Incubation?
binding?

Answer 9

IgM ab lattice w/o additional rgnt
Direct test
Centrifuge brings reactants close
RT
Plasma 2:1 cells 3% sol
37C
(Compliment and IgG ab more likely to bind at body temp)

Question 10

When is the enhancement added in the coombs test?

Answer 10

added after immediate spin and before 37C incubation

this reduces zeta potential
reduces steric hinderance

Question 11

Incomplete agglutinins

Answer 11

usually refers to IgG ab non reactive at RT

Question 12

Coombs

Answer 12

researcher who proposed antiglobulin test method

Question 13

Tube Test

Answer 13

classic serological method preformed with glass tubes

Question 14

AHG

Answer 14

anti human globulin

Question 15

IAT

Answer 15

indirect antiglobulin test

Question 16

Enhanced ab detection

Answer 16

test method requireing more steps than immediate spin to see macroscopic agglutination

Question 17

Briefly cover how the immediate spin test looks

Answer 17

mix cells and plasma
centrifuge
observe agglutination

Question 18

Describe how the coombs test is done

Answer 18

mix cells and plasma
incubate 30-60 min
wash away non attached ab
add anti-ab “coombs rgn”
centrifuge

Question 19

What happens if non attached antibodies are improperly washed?

Answer 19

false negatives

Question 20

If antibody didnt attach to ag at incubation?

Answer 20

add red cells and plasma
incubate
wash
add anti-ab
centrifuge and read for agg
add check cells to agglut w anti-ab

Question 21

Main keys of check cells

Answer 21

have anti-IgG coating the cell
if system isnt washed correctly, anti-ab will attach to ab left over
no binding sites left for binding w check cells

Question 22

How do you confirm a negative reaction in the wash of the coombs test?

Answer 22

check cells

Question 23

What are the 3 possible outcomes of a coombs test

Answer 23

all pos/all neg/ or invalid if check cells are negative

Question 24

When is there an invalid test in coombs?

Answer 24

When the ab in the system is bound to the rgnt not the red cell, causing negative rxn in the check cells

START OVER

Question 25

False negatives in Coombs No agg in AHG exc incor left No agg in CC left ov insuff iinsuff

Answer 25

looks like no agglut in AHG step aged cells excessive washing incorrect centrifuge left over wash dilutes No agglut w check cells left over ab abound check cells insuff washing insuff incubation

Question 26

False positives in coombs test agg in AHG p f ab b develop

Answer 26

looks like agg with AHG step proteins in plasma fibrin strands ab attached in vivo to cells (DAT) bacterial contam pt devel ab to preservative

Question 27

Standardization cell washer monoclonal rgnt gel test solid phase

Answer 27

cell washer: wash step for vol and number of washes, reduces false negatives Monoclonal: binding site, reduces false pos Gel: test and rgnt vol, centrif, removes wash step Solid phase: test/rgnt vol, test ag, interpret

Question 28

LISS

Answer 28

Reduce zeta potential

Question 29

PEG

Answer 29

remove water vol

Question 30

Albumin

Answer 30

increase collisions

Question 31

Bromelin

Answer 31

reduce steric hindrance

Question 32

Polybrene

Answer 32

polymer nonspecific binding

Question 33

Describe gel automated method

Answer 33

precipitation through polyacrylamide gel matrix add red cells and plasma incubate no wash centrifuge no check cells

Question 34

Rouleax looks like what kind of rxn on gel?

Answer 34

looks like 2+ rxn

Question 35

Describe solid phase automated method

Answer 35

adherence of ab to rbc ag on solid substance prone to have false +

Question 36

If gel is available, why tube methods?

Answer 36

Trouble shooting - everyone goes back to tube ref lab method - gold standard tube method is most clinically relevant harder but cheaper

Question 37

When do you do coombs testing? with w sb cross what kind of test? cells in?

Answer 37

with weak D, ab screen, extended cross match, autocontrol Indirect test what has attached to the cell in VITRO

Question 38

Describe direct testing

Answer 38

what is on the cell in the body In VIVO is there already ab on the cell?

Question 39

Requirements for finding compatible blood pt will rx poor if? females? pt with clin sign ab?

Answer 39

pt will react poorly if given incompatible blood females of childbearing age should not be exposed to Rh incomp blood if pt has made clinically sign immune stim ab, they should recive ag neg blood

Question 40

"Blood type" and test

Answer 40

ABO type + Rh type on pt red cells (IS)

Question 41

"Ab screen" and test

Answer 41

test for the presence of clinc sign imm stim ab in pt plasma (IAT)

Question 42

"type and cross match"

Answer 42

ABO + RH+ AB screen + sero between donor cell and pt plasma (IS-IAT)

Question 43

"extended crossmatch" sero test between

Answer 43

sero test between donor cell and pt plasma util at body temp (IAT ONLY)

Question 44

'Ab ID" logical problem utilizing

Answer 44

logical problem utilizing known cells and pt plasma to ID source of unexcpet reactivity in ab screen (IAT)

Question 45

"coombs test"

Answer 45

test for ab attached to cells in VIVO (DAT)

Question 46

Reverse cells

Answer 46

mixture of donors, blood type A or B in preservative (no plasma)

Question 47

Screen cells

Answer 47

single donor type O known ag in preservative no plasma

Question 48

Antigram sheet

Answer 48

package insert that lists all interpretations of ag for screen cells

Question 49

AB screen

Answer 49

each cell represents a single donor (no mf) if any cells in screen rx + more work needs to be done and pt will need extended crossmatch to determ compatibil w donor blood