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IMMUNOHEMA OFFSEM PM > TOPIC 3 - P2 > Flashcards

TOPIC 3 - P2 Flashcards

1
Q

(Direct Antiglobulin Test)

detects in vivo sensitization of RBCs with _____ or __________

A

IgG or complement components

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2
Q

(Direct Antiglobulin Test)

conditions that can result in in vivo coating of RBCs with antibody or complement are:

A
  • Hemolytic disease of the newborn (HDN)
  • Hemolytic transfusion reaction (HTR)
  • Autoimmune and drug-induced hemolytic anemia (AIHA)
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3
Q

not a required test in routine pretransfusion protocols

A

(Direct Antiglobulin Test)

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4
Q

(Direct Antiglobulin Test)

Positive results are monitored by a _______ using monospecific anti-IgG and anti-C3d to determine the specific type of protein sensitizing the cell.

A

DAT panel ;

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5
Q

(Direct Antiglobulin Test)

In an effort to save valuable tech time, some institutions run ______ and ______ reagents at one time as well as a _______

The ________ serves to detect _______ of cells or reactions occurring without the addition of AHG reagents.

A

polyspecific and monospecific ; saline control

  • saline control ; spontaneous agglutination
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6
Q

(Direct Antiglobulin Test)

In the investigation of ______, protein sensitizing the newborn RBCs is presumed to be _______.

A

HDN ; maternal IgG

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7
Q

(Direct Antiglobulin Test)

other techniques can be used to remove antibody from the patient’s RBCs these includes _________, __________, and _________

A

chloroquine diphosphate, EDTA-glycine, and murine monoclonal antibodies

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8
Q

(Evaluation of a Positive DAT)

AABB Technical Manual states that “__________”

A

a positive DAT alone is not diagnostic

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9
Q

(Evaluation of a Positive DAT)

Interpreting the significance of a positive DAT requires knowledge of the _________, _________, ________, and _________

A

patient’s diagnosis, pregnancy, drug therapy, and recent transfusion history

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10
Q

(DAT)

CLINICAL APPLICATION:
HDN
HTR
AIHA

A

IN VIVO SENSITIZATION:
Maternal antibody coating fetal RBCs
Recipient antibody coating donor RBCs
Autoantibody coating individual’s RBCs

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11
Q

DAT PANEL: PATTERNS OF REACTIVITY IN AUTOIMMUNE HEMOLYTIC ANEMIA

Type of AIHA: WAIHA

A

Anti-IgG: +
Anti-C3d: +

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12
Q

DAT PANEL: PATTERNS OF REACTIVITY IN AUTOIMMUNE HEMOLYTIC ANEMIA

Type of AIHA: WAIHA

A

Anti-IgG: +
Anti-C3d: -

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13
Q

DAT PANEL: PATTERNS OF REACTIVITY IN AUTOIMMUNE HEMOLYTIC ANEMIA

Type of AIHA: CAS; PCH, WAIHA

A

Anti-IgG: -
Anti-C3d: +

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14
Q

DAT PANEL: PATTERNS OF REACTIVITY IN AUTOIMMUNE HEMOLYTIC ANEMIA

Type of AIHA: MIXED-TYPE AIHA (cold and warm)

A

Anti-IgG: +
Anti-C3d: +

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15
Q

IN VIVO PHENOMENA ASSOCIATED WITH A POSITIVE DAT (TRANSFUSION)

CONDITION 1. ???

CAUSE 1: Alloantibodies in the recipient of a recent transfusion that react with antigen on donor RBC

A

Recipient alloantibody and donor antigen

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16
Q

IN VIVO PHENOMENA ASSOCIATED WITH A POSITIVE DAT (TRANSFUSION)

CONDITION 2. ???

CAUSE 2: Antibodies present in donor plasma that react with antigen on a transfusion recipient’s RBCs

A

Donor antibody and recipient antigen

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17
Q

IN VIVO PHENOMENA ASSOCIATED WITH A POSITIVE DAT (DRUG INDUCED)

CONDITION 1: ???

CAUSE 1: drug binds covalently to membrane proteins and stimulates hapten-dependent Ab

A

Type 1 (hapten-dependent Ab)

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18
Q

IN VIVO PHENOMENA ASSOCIATED WITH A POSITIVE DAT (DRUG INDUCED)

CONDITION 2: ?????

CAUSE 2: Drug induces autoantibody specific for RBC membrane proteins through unknown mechanism; Ab reacts with normal RBCs in the absence of drug.

A

Type 2/II (autoantibody)

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18
Q

IN VIVO PHENOMENA ASSOCIATED WITH A POSITIVE DAT (DRUG INDUCED)

CONDITION 3: ?????

CAUSE 3: Drug induces Ab that binds to RBC only when drug is present in soluble form, unknown mech; Ab reacts with normal RBCs when soluble drugs is present.

A

Type 3/III (drug-dependent Ab)

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19
Q

IN VIVO PHENOMENA ASSOCIATED WITH A POSITIVE DAT (AUTOIMMUNE HEMOLYTIC ANEMIA)

CONDITION 1: ????
CAUSE 1: autoantibody reacts with patient’s RBCs in vivo

A

WAIHA (IgG or/and C3)

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20
Q

IN VIVO PHENOMENA ASSOCIATED WITH A POSITIVE DAT (AUTOIMMUNE HEMOLYTIC ANEMIA)

CONDITION 2: ???
CAUSE 2: Cold-reactive IgM autoagglutinin binds to RBCs in peripheral circulation (32 deg celsius). IgM binds complement as RBCs return to warmer parts of circulation; IgM dissociates, leaving RBCs coated only with complement.

A

CAS (C3)

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21
Q

IN VIVO PHENOMENA ASSOCIATED WITH A POSITIVE DAT (AUTOIMMUNE HEMOLYTIC ANEMIA)

CONDITION 3: ????
CAUSE 3: The IgG autoantibody reacts with RBCs in colder parts of the body, causes complement to be bound irreversibly to RBCs, and then elutes at warmer temperature.

A

PCH (IgC)

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22
Q

IN VIVO PHENOMENA ASSOCIATED WITH A POSITIVE DAT (HDN)

CONDITION 1: ????
CAUSE 1: Maternal IgG alloantibody, specific for fetal antigen, coats fetal RBCs. DAT is reactive with anti-IgG

A

MATERNAL ALLOANTIBODY CROSSES PLACENTA (IgG)

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23
Q

IN VIVO PHENOMENA ASSOCIATED WITH A POSITIVE DAT (MISCELLANEOUS)

CONDITION 1: ?????

CAUSE 1: Heterophile antibodies that are present in ALG or ATG coat recipient’s RBCs. High levels of protein causing red cells to spontaneously agglutinate.

A

ABSORBED PROTEINS; ADMINISTRATION OF EQUINE PREPARATIONS OF ANTILYMPHOCYTE GLOBULIN AND ANTITHYMOCYTE GLOBULIN

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24
Q

IN VIVO PHENOMENA ASSOCIATED WITH A POSITIVE DAT (MISCELLANEOUS)

CONDITION 2: ????

CAUSE 2: Non-antibody-mediated binding of immunoglobulin to RBCs in patients with hypergammaglobulinemia

A

ADMINISTRATION OF HIGH-DOSE IV GAMMA GLOBULIN AND HYPERGAMMAGLOBULINEMIA

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25
Q

performed to determine in vitro sensitization of RBCs

A

Indirect Antiglobulin Test

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26
Q

Indirect Antiglobulin Test is used in the following situations:

A
  • Detection of incomplete (nonagglutinating) antibodies to potential donor RBCs (compatibility testing) or to screening cells (antibody screen) in serum
  • Determination of RBC phenotype using known antisera (e.g., Kell typing, weak D testing)
  • Titration of incomplete antibodies
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27
Q

(Indirect Antiglobulin Test)

APPLICATION: Antibody detection
TEST:
IN VITRO SENSITIZATION: Recipient antubody reacting with donor cells

A

Compatibility Testing

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28
Q

(Indirect Antiglobulin Test)

APPLICATION: —-
TEST:
IN VITRO SENSITIZATION: Antibody reacting with screening cells

A

Antibody screening (TEST)

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29
Q

(Indirect Antiglobulin Test)

APPLICATION: Antibody identification
TEST:
IN VITRO SENSITIZATION: Antibody reacting with panel cells

A

Antibody panel

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30
Q

(Indirect Antiglobulin Test)

APPLICATION: Antibody titration
TEST:
IN VITRO SENSITIZATION: Antibody and selected Rh cells

A

Rh antibody titer

31
Q

(Indirect Antiglobulin Test)

APPLICATION: RBC phenotype
TEST:
IN VITRO SENSITIZATION: Specific antisera + RBCs to detect antigen

A

RBC antigen detection (ex: weak D, K, Fy)

32
Q

TASKS AND PURPOSES OF THE INDIRECT ANTIGLOBULIN TEST

TASK: Incubate RBCs with antisera
PURPOSE?

A

Allow time for antibody molecule attachment to RBC antigen

33
Q

TASKS AND PURPOSES OF THE INDIRECT ANTIGLOBULIN TEST

TASK: Perform a minimum of three saline washes
PURPOSE?

A

Removes free globulin molecules

34
Q

TASKS AND PURPOSES OF THE INDIRECT ANTIGLOBULIN TEST

TASK: Add antiglobulin reagent
PURPOSE?

A

Forms RBC agglutinates (RBC Ag. + Ab + anti-IgG)

35
Q

TASKS AND PURPOSES OF THE INDIRECT ANTIGLOBULIN TEST

TASK: Centrifuge
PURPOSE?

A

Accelerates agglutination by bringing cells closer together

36
Q

TASKS AND PURPOSES OF THE INDIRECT ANTIGLOBULIN TEST

TASK: Examine for agglutination
PURPOSE?

A

Interprets test as positive or negative

37
Q

TASKS AND PURPOSES OF THE INDIRECT ANTIGLOBULIN TEST

TASK: Grade agglutination reactions
PURPOSE?

A

Determine the strength of reaction

38
Q

TASKS AND PURPOSES OF THE INDIRECT ANTIGLOBULIN TEST

TASK: Add antibody-coated RBCs to negative reactions
PURPOSE?

A

Checks for neutralization of antisera by free globulin molecules (Coombs’ control cells are D-positive RBCs coated with anti-D)

39
Q

(Factors Affecting the Antiglobulin Test)

_______ can detect a level of 100 to 500 IgG molecules per RBC and 400 to 1,100 molecules of C3d per RBC

A

DAT

40
Q

(Factors Affecting the Antiglobulin Test)

_____: must be between 100 and 200 IgG or C3 molecules on the cell to obtain a positive reaction

A

IAT

41
Q

Factors Affecting the Antiglobulin Test

A
  1. Ratio of serum to cells
  2. Reaction Medium
  3. Temperature
  4. Incubation Time
  5. Washing of RBCs
  6. Saline for Washing
  7. Addition of AHG
  8. Centrifugation for reading
42
Q

(Factors Affecting the Antiglobulin Test)

  1. Ratio of serum to cells

Increasing the ratio of serum to cells increases the ________

A

sensitivity of the test system

43
Q

(Factors Affecting the Antiglobulin Test)

  1. Ratio of serum to cells

minimum ratio of ___ should be the target; using ___ drops of serum and 1 drop of a _______

A

40:1 ; 2 ; 5% suspension of cells

44
Q

(Factors Affecting the Antiglobulin Test)

  1. Ratio of serum to cells

When using cells suspended in saline, it is often advantageous to increase the ratio of serum to cells to detect weak antibodies (e.g., ____ drops of serum with 1 drop of a ______; ratio of _____)

A

4 ; 3% cell suspension ; 133:1

45
Q

(Factors Affecting the Antiglobulin Test)

  1. Reaction Medium

A. ALBUMIN

macromolecules of albumin allow ________ to come into closer contact with each other so that aggregation occur

A

antibody-coated cells

46
Q

(Factors Affecting the Antiglobulin Test)

  1. Reaction Medium

A. ALBUMIN

1965: ________ reported on the creased sensitivity of the IAT if albumin was incorporated

2 drops of _____, 2 drops of ________, and 1 drop of _______, _____ minutes of incubation

A

Stroup and MacIlroy

serum ; 22% bovine albumin ; 3% to 5% cells ; 30

47
Q

(Factors Affecting the Antiglobulin Test)

  1. Reaction Medium

A. ALBUMIN

_______: showed that an albumin technique may miss several clinically significant antibodies

A

Petz and coworkers

48
Q

(Factors Affecting the Antiglobulin Test)

  1. Reaction Medium

B. LISS (low ionic strength solution)

enhance antibody uptake and allow incubation times to be decreased—from 30 to 60 minutes to _______—by reducing the _______ surrounding an RBC

A

10 to 15 minutes ; zeta potential

49
Q

(Factors Affecting the Antiglobulin Test)

  1. Reaction Medium
    B. LISS (low ionic strength solution)

Introduced by ________

A

Low and Messeter

50
Q

(Factors Affecting the Antiglobulin Test)

  1. Reaction Medium
    B. LISS (low ionic strength solution)

___________ showed that optimum reaction conditions were obtained using 2 drops of serum and 2 drops of a 3% suspension of cells in LISS

Increasing the serum-to-cell ratio increased the ________ of the reaction mixture, leading to a decrease in _______ of the test

A

Moore and Mollison

ionic strength ; sensitivity

51
Q

(Factors Affecting the Antiglobulin Test)

  1. Reaction Medium
    C. Polyethylene glycol (PEG)

water-soluble linear _______

A

polymer

52
Q

(Factors Affecting the Antiglobulin Test)

  1. Reaction Medium
    C. Polyethylene glycol (PEG)

used as an additive to increase antibody uptake by removal of _______ surrounding the RBC (the water of hydration theory), thereby effectively concentrating antibody

A

water molecules

53
Q

(Factors Affecting the Antiglobulin Test)

  1. Reaction Medium
    C. Polyethylene glycol (PEG)

Anti-IgG is the AHG reagent of choice to avoid _______

A

false-positive reactions

54
Q

(Factors Affecting the Antiglobulin Test)

  1. Reaction Medium
    C. Polyethylene glycol (PEG)

_____ incubation in the IAT is omitted

A

37°C

55
Q

(Factors Affecting the Antiglobulin Test)

  1. Reaction Medium
    C. Polyethylene glycol (PEG)

increases the detection of ________ while decreasing detection of _________

A

clinically significant antibodies ; clinically insignificant antibodies

56
Q

(Factors Affecting the Antiglobulin Test)

  1. Temperature

______: usual incubation temperature for the IAT; optimum temperature for complement activation; where the rate of reaction for the majority of IgG antibodies is optimal

A

37°C

57
Q

(Factors Affecting the Antiglobulin Test)

  1. Incubation Time

cells suspended in saline: _____________
majority of clinically significant antibodies can be detected after ________ of incubation

A

30 and 120 minutes incubation
30 minutes

58
Q

(Factors Affecting the Antiglobulin Test)

  1. Incubation Time

________: 10 to 15 minutes incubation

A

LISS or PEG: 10 to 15 minutes incubation

59
Q

(Factors Affecting the Antiglobulin Test)

  1. Washing of RBCs

RBCs must be saline-washed a minimum of _______ before adding AHG reagent

  • removes __________
A

three times ; free unbound serum globulins

60
Q

(Factors Affecting the Antiglobulin Test)

  1. Washing of RBCs

Inadequate washing may result in a ______

  • neutralization of the AHG reagent by _________
A

false-negative reaction ; residual unbound serum globulins

61
Q

(Factors Affecting the Antiglobulin Test)

  1. Washing of RBCs

Washing should be performed immediately after being removed from the incubator in as short a time as possible

  • to minimize the ____________-
A

elution of low-affinity antibodies

62
Q

(Factors Affecting the Antiglobulin Test)

  1. Washing of RBCs

All saline should be discarded after the final wash, because residual saline ________

  • decreases the ________ of the test
A

dilutes the AHG reagent ; sensitivity

63
Q

(Factors Affecting the Antiglobulin Test)

  1. Saline for washing

should be fresh or buffered to a pH of _____ to ______

A

7.2 to 7.4

64
Q

(Factors Affecting the Antiglobulin Test)

  1. Saline for washing

________: decrease in pH, which may increase the rate of antibody elution during the washing process, yielding a false-negative result

A

Saline stored for long periods in plastic containers

65
Q

(Factors Affecting the Antiglobulin Test)

  1. Addition of AHG

added to the cells immediately after washing to minimize ___________ and subsequently __________

A

the chance of antibody eluting from the cell ; neutralizing the AHG reagent

66
Q

(Factors Affecting the Antiglobulin Test)

  1. Centrifugation for Reading

Centrifugation of the cell pellet for reading of hemagglutination along with the method used for resuspending the cells: __________-

A

crucial step in the technique

67
Q

(Factors Affecting the Antiglobulin Test)

  1. Centrifugation for Reading

_____ RCFs for _____ seconds (Harmening)

A

500 ; 15 to 20

68
Q

(Factors Affecting the Antiglobulin Test)

  1. Centrifugation for Reading

____ RCFs for ____ seconds(CBER)

A

1,000 ; 20

69
Q

(Factors Affecting the Antiglobulin Test)

  1. Centrifugation for Reading

use of _____ RCFs yields more sensitive results

A

higher

70
Q

(Factors Affecting the Antiglobulin Test)

  1. Centrifugation for Reading

weak false-positive results because of ______

A

inadequate resuspension

71
Q

(Factors Affecting the Antiglobulin Test)

  1. Centrifugation for Reading

false negative result if resuspension is ______

A

too vigorous

72
Q

(Sources of Error)

An _________ should be used to collect blood samples for the DAT to avoid the in vitro complement attachment associated with refrigerated clotted specimens

A

anticoagulant (EDTA)

73
Q

(Sources of Error)

All negative antiglobulin test reactions must be checked by the addition of ________

should demonstrate _______ of these RBCs with the anti-IgG in the AHG reagent

A

IgG-sensitized cells (Check cells) ; hemagglutination

74
Q

(Sources of Error)

If no hemagglutination follows the addition of IgG-coated RBCs, the test result is ________ and ________

most common technical errors: _______, _______, and _________

A

invalid and the test must be repeated ;

inadequate washing, nonreactive AHG reagent, and failure to add AHG reagent

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