Microsoft word - ct.004 _rev 2.0_.doc

Rh Typing Problem Solving

1.0 Principle
2.0 Scope and Related Policies
2.1. Rh typing should be investigated when: • The Rh control test is positive. 9.1 (F3.2), 9.2 (10.4.4) • Results are weak or 1+ positive with commercial anti-D reagent. Microscopic readings should only be done if mixed field agglutination is suspected.

• Rh typing discrepancies are found between current and previous results. 9.1 (G5.3), 9.2 (10.4.6) 2.2. Previous transfusion records shall be reviewed. Previous results shall be compared with current results. 9.1 (G5.2; G5.3), 9.2 (10.4.6) 2.3. All reagents shall be used and controlled according to the supplier’s recommendations and procedures. 9.1 (F1.1), 9.2 (8.1.2) 2.4. If an Rh typing problem is detected and transfusion is necessary before resolution, Rh negative blood products should be issued
until the problem is resolved, especially for children and women of
child-bearing age. 9.1 (G9.2.5), 9.2 (

2.5. False positive reactions with anti-D (control negative) may be caused by the presence of antibodies to antigens of low frequency in antisera of human origin. Repeating the Rh typing with monoclonal reagents should resolve the problem. Monoclonal reagents do not have contaminating antibodies. 3.0 Specimens
4.0 Materials
Technical Resource Manual for Hospital Transfusion Services Rh Typing Problem Solving
5.0 Quality Control/Management
5.1. See QCAI.001 - Quality Control of Reagent Red Cells and Antisera. 5.2. False positive results in the Rh control may occur in any saline reactive test system if the serum contains autoagglutinins or the red cells are tested unwashed when rouleaux is present. A negative reaction with anti-A or anti-B usually rules out spontaneous agglutination. 5.2.2. An Rh control is suggested for all Rh tests. If this is not desired a control shall be performed if the recipient types as AB Rh positive. A control is not required when donor units are confirmed. 5.2.3. If a commercial control for low protein anti-D (monoclonal/ polyclonal blended) is not available, autologous serum/plasma or a 6–8% bovine albumin control may be used to control Rh typing. 5.2.4. A DAT should be performed as a control for selected samples (e.g., cord, neonatal and samples demonstrating autoagglutination) in order to rule out false negative Rh typing due to blocked antigens. 6.0 Procedure
6.1. Recheck suitability of all specimen(s). See PA.002 - Determining
Specimen Suitability. If the specimen is unsuitable, collect another specimen. 6.2. If there is any doubt about the identity or the quality of the specimen, collect a new specimen and repeat testing. 6.3. Check the label on the reagent vial(s) to ensure that the correct 6.4. Recheck the reagent’s appearance for possible contamination.
Use a new reagent vial if the current vial appears to be contaminated. Technical Resource Manual for Hospital Transfusion Services Rh Typing Problem Solving
6.5. Prepare a new 3% recipient washed red cell suspension.
6.6. Repeat the Rh typing on the new 3% recipient cell suspension. 6.7. If problem is resolved, see procedural note 8.1 and proceed to step 6.8. If problem is unresolved and transfusion is required STAT, select Obtain and record the recipient’s diagnosis and transfusion/obstetrical history. Identify the Rh problem and proceed as follows: • If the reaction with anti-D is weak or 1+ positive and the control is negative, go to step 6.10. • If the control is positive, go to step 6.11. • If the result is discrepant (the previous Rh typing does not agree with the current test result) go to step 6.12. 6.10. If the reaction with anti-D is weakly positive or 1+ and the control
and read the tubes microscopically and look for mixed field agglutination. See procedural note 8.2. If the recipient has been transfused in the last 3 months with blood of a different Rh group, record the historical grouping on the current record with an explanation for the discrepancy (e.g. Recipient transfused with ___ units of Rh positive RBC on ____). If a large fetomaternal bleed has occurred and the mother’s specimen tests weakly positive with Anti-D, record the historical Rh type with an explanation for the If the recipient has not been transfused in the last 3 months, perform a weak D typing test. See RT.003 – Technical Resource Manual for Hospital Transfusion Services Rh Typing Problem Solving
If the discrepancy has not been resolved see 6.11. If the control is positive:
6.11.1. If the anti-D and Rh control are high-protein reagents, the Rh typing should be repeated using a low-protein reagent such as monoclonal anti-D and control. If the low-protein control is still positive, the antibody coating the cells should be removed before repeating the Rh typing. cells 4 times with saline (if cold agglutinin is suspected, use 37o C saline to wash the cells). See PA.005 - Cell Washing Automated and Manual. • Repeat the Rh typing on the 3% washed cell
• If the control is negative when using a 3% washed
cell suspension, report the Rh typing. See procedural note 8.3 for possible causes. Proceed to step 6.13. 6.11.3. If the control is still positive using washed red cells, perform a Direct Antiglobulin Test (DAT). See RT.004 - Direct Antiglobulin Test. If the DAT is positive, refer the specimen out for or perform a chloroquine treatment (or alternative method) to remove the antibody coating the cells. Follow the manufacturer’s directions for chloroquine treatment of cells. • Repeat the Rh testing using chloroquine- • If the discrepancy is resolved, report the Rh type with a note that the test was done using chloroquine-treated cells. • If the discrepancy is not resolved, send the specimen to a reference lab for additional 6.12. If the results are discrepant and the previous or historical Rh
typing does not agree with the current test result: Technical Resource Manual for Hospital Transfusion Services Rh Typing Problem Solving
6.12.1. Review recipient’s medical history: • transfusion - If the recipient was transfused in the last 3 months with blood of a different Rh type, follow steps 6.10.1 and 6.10.2. Recipient not
Then Consider
or historical specimen. Review the historical test results, if possible. disease state. Check diagnosis. Consider adsorption and elution with anti-D. See step 8.4.2. Then Consider

Result on recollected
6.13. Complete an incident report as per laboratory policy and submit to Technical Resource Manual for Hospital Transfusion Services Rh Typing Problem Solving
6.14. Report the process and the conclusion of the investigation on the 6.15. When the procedure is complete, perform a clerical check. 6.16. Sign or initial and record the completion time and date on the request form. Omit this step if these are captured by a computer 7.0 Reporting
• All procedures that were used to resolve the Rh typing problem (e.g., washing the cell suspension, use of different manufacturers and/or lot number of anti-D, etc.) 7.2. When a result is discrepant from a previous Rh typing: • A recipient previously typed as Rh negative is found to be Rh positive after investigation, the recipient Rh type should be reported as (for example): “Recipient types as Rh positive. Previous Rh negative reported could have been due to undetected weak expression of the D antigen, technical or clerical error on the previous specimen”.
• A recipient previously typed as Rh positive is found to be Rh
negative after investigation, the recipient Rh type should be reported as (for example): “Recipient red cells type as Rh negative. Previous Rh positive reported could have been due to a technical or clerical error on the previous specimen.” • The discrepancy is not resolved. Refer to supervisor and/or pathologist. Report as “Rh cannot be determined at this time. If red cells are required, Rh negative red cells will be issued". 8.0 Procedural Notes
8.1. If the discrepancy has been resolved, one of the following could • Antiserum not added, or wrong one used Technical Resource Manual for Hospital Transfusion Services Rh Typing Problem Solving
• Centrifugation insufficient or excessive • Tube shaken too vigorously and small agglutinates were • Failure to resuspend entire cell button • Reading microscopically when antisera instructions indicate 8.2. Possible causes for mixed field agglutination: • Recent transfusion with different Rh donor unit • Unusual Rh phenotype that may or may not be associated with production of Rh alloantibodies

• Dual red cell population as result of bone marrow transplant • Genetic anomalies such as dispermy and chimerism. 8.3. Positive control may be due to:
• Rouleaux when using unwashed cell suspensions • Positive Direct Antiglobulin Test (DAT). 8.4. False negative reactions with anti-D may occur: When the cells are coated with an antibody specific for
the antigen being tested. Example: when the D antigen
sites on fetal cells are coated with the maternal anti-D,
the neonate’s red cells may fail to react with the anti-D
and therefore type as Rh negative.

In disease states that result in loss of antigen. Recipients who were known to be D positive have been studied during their illness and were found to be D negative by “weak D” typing test. Technical Resource Manual for Hospital Transfusion Services Rh Typing Problem Solving

9.0 References
9.1. Canadian Society for Transfusion Medicine. Standards for hospital transfusion services, version 1. Ottawa: Canadian Society for Transfusion Medicine, 2004: F1.1, F3.2, G5.2, G5.3, G9.2.5. 9.2. Canadian Standards Association. Blood and blood components (CAN/CSA Z902-04). Mississauga, Ontario: Canadian Standards Association, 2004: 8.1.2, 10.4.4, 10.4.6, Facility endorsement if guideline is used as a Standard Operating
Procedure (SOP)
Change Log
Change Description
Effective Date

Revision 1
5.2.1: corrected numbering
6.10.4: added step for unresolved discrepancy
last bullet - added step to see 7.4 for unresolved discrepancy
7.4: added step for reporting unresolved discrepancy
Updated reference
Revision 2
Changed “patient” to “recipient” in all cases where applicable
2.4:Added “especially for children and women of child-bearing age”
5.0: Updated title
9.0: Deleted references 9.2–9.4; added new reference 9.2.
Technical Resource Manual for Hospital Transfusion Services


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