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Investigation of Positive Direct Antiglobulin Test (DAT)

Guideline: CT.005(Rev2.0) Guideline Effective Date: DDMMMYY Revision Date: 01JAN05

1.0

Principle

To investigate the possible causes of a positive Direct Antiglobulin Test (DAT). Investigation of a positive DAT will assist in establishing a diagnosis and/or in transfusion therapy.

2.0

Scope and Related Policies

2.1. There shall be policies and procedures for documentation, reporting, evaluation and follow-up of all transfusion reactions. 9.1 (N1.1), 9.3 (17.1.1) 2.2. If there is insufficient serum/plasma from the neonate, maternal serum may be used for crossmatching and antibody screening. 9.1 (J2.1),

9.3 (10.9.1.5)

2.3. In recipients with previously identified clinically significant antibodies, the antibody identification shall be performed when there is clinical or serological evidence of a new antibody. 9.2 ( 5.13.3.3)

3.0

Specimens

N/A

4.0

Materials

N/A

5.0

Quality Control/Management

N/A

6.0

Procedure

6.1. 6.2. If recipient is a neonate, go to step 6.7. Obtain the following information: · · Medication history Transfusion in the past 3 months.

It may be necessary to ask the recipient, the recipient's family, the attending nurse and/or the physician to obtain an accurate history.

Technical Resource Manual for Hospital Transfusion Services

CT.005(Rev2.0) Page 1 of 8

Investigation of Positive Direct Antiglobulin Test (DAT)

Guideline: CT.005(Rev2.0) Guideline Effective Date: DDMMMYY Revision Date: 01JAN05

6.3.

Consider the possibility of a delayed hemolytic transfusion reaction if the recipient has been transfused in the past 3 months (or if the history of transfusion is unsure or unknown). See procedural note 8.1. 6.3.1. Find out where the transfusion occurred and contact the hospital to inquire about the pretransfusion antibody screen and DAT status.

6.3.2. Perform an antibody screen or a group and screen on the current specimen. 6.3.2.1. If the antibody screen is positive, perform an antibody identification. See CT.007 - Antibody Identification of Warm Reactive Antibodies.

6.3.2.2. If all cells (screening and panel) are positive, consider performing an allogeneic red cell adsorption or sending specimens to a reference laboratory for investigation. See procedural note 8.2. 6.3.3. Prepare an eluate or send the specimen to a reference laboratory for an elution. See CT.010 ­ Acid Elution. An elution may be omitted if all the criteria in procedural note 8.3 are met or if directed by a laboratory physician.

6.4. 6.5.

If the recipient has not been transfused in the past 3 months, an eluate is not required. See procedural note 8.4. When the DAT is positive with anti-C3 and negative with anti IgG, Cold Agglutinin Disease (CAD) or Paroxysmal Cold Hemoglobinuria (PCH) should be considered. Usually results are strongly positive in these cases. If these clinical conditions are suspected, the following test(s) may be performed: 6.5.1. 6.5.2. Cold agglutinin screen to confirm CAD. Donath-Landsteiner (D-L) test to confirm PCH.

6.6.

Whether or not the recipient has been transfused, it may be necessary to evaluate the recipient for evidence of hemolysis caused by premature destruction of the coated red cells. This will depend on clinical presentation and investigation and additional testing shall be requested by a physician.

Technical Resource Manual for Hospital Transfusion Services

CT.005(Rev2.0) Page 2 of 8

Investigation of Positive Direct Antiglobulin Test (DAT)

Guideline: CT.005(Rev2.0) Guideline Effective Date: DDMMMYY Revision Date: 01JAN05

6.6.1.

The following tests may be ordered by a pathologist or designate: · · · Hemoglobin Red cell morphology Additional tests such as a reticulocyte count, bilirubin, etc. See procedural note 8.5.

6.6.2.

If the recipient does not show evidence of hemolysis as determined by a physician, or if a transfusion is not anticipated, see procedural note 8.4 and report as in 7.3 ­ Reporting. Refer any request for transfusion to a pathologist or designate if the recipient shows evidence of hemolysis because of the risks of aggravating the hemolysis and/or alloimmunizing the recipient. See procedural notes 8.6 and 8.7.

6.6.3.

6.7.

If recipient is a neonate and has a positive DAT or is jaundiced and has a negative DAT, the physician should consider hemolytic disease of the newborn (HDN). 6.7.1. Perform the following tests: · · ABO grouping on the maternal and neonatal specimens Antibody screen on the maternal specimen. If the antibody screen is positive, perform antibody identification. See CT.007 - Antibody Identification of Warm Reactive Antibodies.

6.7.2. When the maternal antibody suspected of causing the positive DAT is other than ABO, type the neonatal cells for the corresponding antigen(s). See CT.009 - Antigen Typing - Direct and Indirect Agglutination. Antisera for phenotyping should not be used if it requires IAT testing (use monoclonal antisera whenever possible). The neonate's cells should be positive for the antigen if the maternal antibody is causing the positive DAT. See procedural note 8.8.

Technical Resource Manual for Hospital Transfusion Services

CT.005(Rev2.0) Page 3 of 8

Investigation of Positive Direct Antiglobulin Test (DAT)

Guideline: CT.005(Rev2.0) Guideline Effective Date: DDMMMYY Revision Date: 01JAN05

6.7.3. If the maternal serum or plasma antibody screen is negative and there is no ABO incompatibility and the neonatal specimen has a positive DAT, consider the possibility of a maternal antibody directed against a low frequency antigen (e.g., anti-Cw, -Kpa, -Jsa, etc.). 6.7.4. Interpret neonatal results.

DAT on neonate Maternal Antibody Status ABO Incompatibility Interpretation Positive DAT probably due to ABO antibody Positive DAT may be due to alloantibody and/or ABO antibody - an elution is required to determine Positive DAT probably due to maternal alloantibody Positive DAT probably due to ABO antibody - an elution is required to determine Positive DAT probably due to maternal alloantibody Although the DAT is negative, HDN due to maternal anti___ (-A or -B) cannot be excluded

Positive Positive

Negative Clinically Significant Antibody Clinically Significant Antibody Inconclusive or Insignificant Antibody Inconclusive Antibody Negative

Yes Yes

Positive Positive Positive Negative

No Yes No Yes

6.7.5.

Report DAT results. See 7.4 - Reporting.

7.0

Reporting

7.1. If the recipient has been transfused in the past 3 months and the elution was not done, report: "Positive DAT. Anti-IgG ____; anti-C3 ___. Elution not done. Recipient has not received blood components in the last 3 weeks and there has been no change in the DAT and antibody screen results since specimen tested on __________ (insert date of testing)." If the recipient has not been transfused in the last 3 months, report: "Positive DAT. Anti-IgG ____; anti-C3 ____. Elution not done because recipient has not been transfused in the last 3 months." If the elution was performed, see CT.010 - Acid Elution - 7.0 Reporting.

CT.005(Rev2.0) Page 4 of 8

7.2.

7.3.

Technical Resource Manual for Hospital Transfusion Services

Investigation of Positive Direct Antiglobulin Test (DAT)

Guideline: CT.005(Rev2.0) Guideline Effective Date: DDMMMYY Revision Date: 01JAN05

7.4.

For neonatal specimens, evaluate the test results and report "Positive DAT probably due to maternal anti-___ ." 7.4.1. If the DAT is negative, the neonate is jaundiced, the maternal antibody screen is negative and neonatal cells are ABO incompatible with the maternal serum/plasma, report: "Although the DAT is negative, HDN due to maternal anti___ (-A or -B) cannot be excluded."

8.0

Procedural Notes

8.1. A positive DAT developing after a recent transfusion should always alert staff to the possibility of a delayed hemolytic transfusion reaction. Recovery of alloantibody in a red cell eluate suggests a hemolytic (delayed) transfusion reaction. The alloantibody may or may not be present in the serum/plasma. When an alloantibody can be demonstrated in the serum/plasma, sometimes a second alloantibody can be found in the eluate. An elution may also be done when the DAT is positive with only complement. Usually warm autoantibodies mimic a multiple complex Rh specificity but autoantibodies against other blood group systems may also be present. Recipients with previous transfusions or pregnancies may have alloantibodies that are masked by the presence of the autoantibody. Adsorption procedures may be required to determine whether an alloantibody is present. If the recipient has been transfused within the past 3 months, but not within the last 3 weeks, an elution may be omitted if all of the following apply: 8.3.1. The DAT was positive on the last specimen tested (i.e., collected more than 3 weeks ago) and was investigated and The positive DAT (i.e., strength of the reaction) on the current specimen is not stronger than the DAT performed on the last specimen and The antibody screen (i.e., strength of the reaction) on the current specimen is the same as on the last specimen.

8.2.

8.3.

8.3.2.

8.3.3.

Note: Clinical circumstances, evaluation of transfusion, test result history and/or specimen history may override the above criteria and elution may be desirable for selected recipients.

Technical Resource Manual for Hospital Transfusion Services CT.005(Rev2.0) Page 5 of 8

Investigation of Positive Direct Antiglobulin Test (DAT)

Guideline: CT.005(Rev2.0) Guideline Effective Date: DDMMMYY Revision Date: 01JAN05

8.4. Approximately 1 in 10,000 individuals have a positive DAT with antiIgG and/or anti-C3. The significance of the positive DAT is unknown. 8.5. A positive direct antiglobulin test may result from: 8.5.1. Alloantibodies in a recipient's circulation reacting with antigens on recently transfused donor cells. 8.5.2. Passively acquired alloantibodies in donor plasma, platelets, or plasma derivatives which react with antigens on the red cells of a transfused recipient (e.g., these antibodies may include anti-A,B, anti-A and/or anti-B, anti-D from Rh Immune Globulin, etc.). 8.5.3. Passively acquired antibodies in a neonate from maternal circulation. 8.5.4. Non-red-cell immunoglobulins associated with red cells in recipients with hypergammaglobulinemia or recipients of high-dose intravenous gammaglobulin. 8.5.5. Autoantibodies to intrinsic red cell antigens. Antibodies directed against certain drugs that bind to red cell membranes (e.g., penicillin, methyldopa, etc.) 8.6. In the presence of a warm autoantibody, transfusion should be avoided if possible and should only be given for severe lifethreatening anemia. Medical treatment ordered by a physician should be the mainstay of treatment when possible (e.g., steroids). 8.7. Serologic investigations should be performed before a transfusion is required. The results of these investigations can be very helpful in selecting appropriate units if transfusion is required. Laboratories should anticipate the possibility of future transfusion requirements and perform these investigations well in advance of transfusion requests. 8.8. When a maternal antibody has been identified as the cause of a positive DAT on a neonatal specimen, exchange transfusion should be anticipated. If the neonate's clinical condition indicates the possibility of exchange transfusion as determined by a physician, the transfusion service should make arrangements to obtain suitable donor units. See CSP.001 ­ Selection of Blood Components for Transfusion.

CT.005(Rev2.0) Page 6 of 8

Technical Resource Manual for Hospital Transfusion Services

Investigation of Positive Direct Antiglobulin Test (DAT)

Guideline: CT.005(Rev2.0) Guideline Effective Date: DDMMMYY Revision Date: 01JAN05

9.0

References

9.1. Canadian Society for Transfusion Medicine. Standards for hospital transfusion services, version 1. Ottawa: Canadian Society for Transfusion Medicine, 2004: J2.1, N1.1. Fridey JL, ed. Standards for blood banks and transfusion services, 22nd ed. Bethesda, MD: American Association of Blood Banks, 2003: 5.13.3.3. Canadian Standards Association. Blood and blood components (CAN/CSA Z902-04). Mississauga, Ontario: Canadian Standards Association, 2004: 10.9.1.5, 17.1.1.

9.2.

9.3.

Facility endorsement if guideline is used as a Standard Operating Procedure (SOP) Approved By: (Senior Management) (Senior Management) Facility effective date: DDMMMYY (Date of implementation)

Technical Resource Manual for Hospital Transfusion Services

CT.005(Rev2.0) Page 7 of 8

Investigation of Positive Direct Antiglobulin Test (DAT)

Guideline: CT.005(Rev2.0) Guideline Effective Date: DDMMMYY Revision Date: 01JAN05

Change Log Change Description

Original Revision 1 Removed steps relating to elution to new guideline - CT.010 - Acid Elution and referenced. Previous 6.5.4: Removed steps regarding follow-up testing if transfusion is required. This information will be in other guidelines (e.g., RT.005, CT.007, etc.). 6.5: Clarified wording. 6.7.2: Added use of antisera not requiring IAT testing. 6.7.4.: Added table for interpretation of neonatal results. Moved information regarding this from reporting section and condensed neonatal reporting. 7.1: Expanded section. Previous 7.3: Deleted - moved to elution (CT.010) procedure. 8.3: Moved information from procedure to procedural notes and referenced to 6.3.3. Added note regarding exceptional circumstances. 9.2: Updated reference. 9.4: Added reference for 3 week rule for no elution required. Revision 2 Changed "patient" to "recipient" in all cases where applicable 5.0: Updated title 9.0: Deleted references 9.3­9.5; Added new reference 9.3.

Effective Date

April 2000

November 2000

January 2005

Technical Resource Manual for Hospital Transfusion Services

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