DAT & IAT/ABO Discrepancies

Question 1

Direct Antiglobulin Test

Answer 1

checking for sensitized RBC’s in vivo

Question 2

Indirect Antiglobulin Test

Answer 2

Checking for sensitized RBC’s in vitro

Question 3

When is the DAT ordered?

Answer 3

Hemolytic disease of the newborn investigation
Transfusion reaction investigation
Immune hemolytic anemia
Drug absorption

Question 4

Weak d investigation
Antibody screen
Antibody identification panel
Crossmatch (if carried out through all phases)

Answer 4

Tests that commonly require an IAT

Question 5

Polyspecific AHG

Answer 5

Contains both anti-IgG and anti-C3d.

Allows the reaction of agglutination to be seen for one or both of these antibodies

Question 6

Monospecific AHG

Answer 6

Individual investigations can be performed to find whether an antibody on the RBC is causing the agglutination or the presence of complement is causing the agglutination

Question 7

How can AHG be inactivated?

Answer 7

Free antibodies that are not attached to the RBC surface

Question 8

What will happen if we have inactivated the AHG before it has an opportunity to interact with the antibodies on the surface of the RBCs?

Answer 8

False negative reaction and Coombs control (check cells) will be negative

Question 9

What does a positive result indicate on the AHG control test (check cells)?

Answer 9

Free AHG was available to combine with the sensitized RBC’s (coombs control reagent) and cause agglutination

Question 10

Do you perform a AHG control test if the tube has agglutination in the AHG phase of the test?

Answer 10

No, the AHG is reacting. We are checking for false negatives using the Coombs control cells

Question 11

What does a negative result indicate on the AHG control test?

Answer 11

We don’t know where the AHG is, invalid test

Question 12

An adult patient is found to have a positive direct antiglobulin test. How would you investigate?

Answer 12

1) check medications

2) elution, then antibody ID if caused by an antibody

Question 13

3 causes of in vivo RBC sensitization

Answer 13

HDN, HTR, and AIHA

Question 14

If a patient has autoimmune hemolytic anemia, what may be found on the patients RBC?

Answer 14

Autoantibody/complement

DAT positive, along with Rh control tube (possibly)

Question 15

Common antibiotics that may absorb onto the RBC membrane that will give a positive DAT result

Answer 15

Penicillins
Cephalosporins
Erythromycin

Question 16

Drugs that may cause an autoantibody mechanism that would give a positive DAT

Answer 16

Methyldopa
Levodopa
Mefinamic acid
Procainamide
Ibuprofen

Question 17

Significance of a positive DAT result

Answer 17

Indicates that antibody has attached to the red cells in vivo

Question 18

What phase does sensitization primarily occur in the IAT?

Answer 18

37 degrees C in vitro

Question 19

Where does sensitization occur in the DAT?

Answer 19

In vivo

Question 20

Zeta potential

Answer 20

Natural repulsion between RBCs. Caused by ionic charge on surface and surrounding RBCs

Question 21

Where does AHG anti-IgG bind on the antibody molecule?

Answer 21

Fc portion

Question 22

Where does AHG anti-complement bind on the complement molecules?

Answer 22

C3d

Question 23

Primary usefulness of albumin

Answer 23

Allows Rh antibodies to appear at 37 phase - is not as broadly reactive as the others

Question 24

Primary usefulness of LISS

Answer 24

Low ionic strength saline has broad use - but reaction’s primarily in AHG phase only of testing

Question 25

Primary usefulness of PEG

Answer 25

Enhancement of reactions, some problems with interpretation, can't read at 37 degree phase

Question 26

Which potentiator increases the affinity of the antibody and antigen instead of lowering the zeta potential?

Answer 26

PEG

Question 27

Which potentiator is used when the interpretation of the results is not done in the 37 degree phase of an IAT?

Answer 27

PEG

Question 28

Order of complement

Answer 28

C1, C4, C2, C3, C5, C6, C7, C8, C9

Question 29

Main purpose of AHG

Answer 29

Attaches to the surface of RBCs and spans the distance between RBCs to cause agglutination

Question 30

Charge on the surface of a red blood cell

Answer 30

Net negative charge surrounding the RBC

Question 31

What specifically is Coombs control (check cells)?

Answer 31

Sensitized RBC's

Question 32

What is the purpose of the AGH control (check cells) test?

Answer 32

Check to see if AHG has bridged between IgG antibodies or if the AHG is still free

Question 33

Why would monospecific AHG (anti-IgG) most likely not react to an anti-M in the AHG phase?

Answer 33

Anti-M is an IgM, cold-reacting antibody

Question 34

Why might polyspecific AHG react with an IgM antibody at the AHG phase and monospecific AHG will not?

Answer 34

Complement may have bound and left over compliment proteins are on the surface of the RBC

Question 35

What would happen in a DAT test if compliment was bound to the RBC's and polyspecific AHG was used?

Answer 35

False positive: compliment and anti-C3d

Question 36

If a patient had an IgM antibody that was a broad thermal range and you were using monospecific AHG, where would the agglutination take place?

Answer 36

Positive IS and 37 phase, negative AHG phase

Question 37

In an indirect or direct AHG test, what is reacting together to cause agglutination in the check cells step?

Answer 37

AHG and check cells

Question 38

If a person had an autoimmune hemolytic anemia caused by an IgG, where would you expect to see a positive result - indirect AHG or direct AHG?

Answer 38

Direct

Question 39

If a person had an IgG autoantibody, why wouldn't you normally see a discrepancy in the reverse testing?

Answer 39

IgG is a warm reacting antibody - it wouldn't bind to its antigen at room temp

Question 40

How do you separate the antigen and antibody in a patient that has a positive DAT if you want to ID the antibody?

Answer 40

Elution

Question 41

Possible outcomes of compliment binding

Answer 41

Hemolysis by lysis of RBC | Destruction of RBC by a macrophage recognizing compliment as an opsonin

Question 42

What class of antibody is most likely identified in the AHG phase of testing?

Answer 42

IgG

Question 43

Which compliment pathway is detected using polyspecific AHG?

Answer 43

Classical