How should I perform and report ABO and Rh blood grouping on an EDTA whole‑blood sample for transfusion, organ transplantation, or pregnancy management?

Blood Grouping Test: Performance and Reporting Standards

Core Testing Requirements

For transfusion, organ transplantation, or pregnancy management, perform ABO and RhD typing on EDTA whole-blood samples using serological methods with both forward (cell) and reverse (serum) typing to confirm ABO group, and always include antibody screening to detect unexpected alloantibodies. 1, 2

Standard Testing Protocol

Forward and Reverse Typing:

• Forward typing detects A, B, and D antigens on the patient's red blood cells using specific antisera 1
• Reverse typing confirms ABO group by detecting expected isoagglutinins (anti-A, anti-B) in the patient's serum 1
• Both methods must agree for valid ABO typing 1

Antibody Screening:

• Perform antibody screening on all samples to detect unexpected alloantibodies that could cause hemolytic transfusion reactions 1, 3
• This is mandatory before any transfusion, transplantation, or pregnancy management 1

Sample Validity and Timing

The 72-Hour Rule:

• Type and screen samples remain valid for 3 calendar days (72 hours) in patients without recent transfusion or pregnancy 4
• If the patient received any transfusion OR was pregnant within the past 3 months, the sample expires after only 72 hours and must be repeated 4
• New antibodies can develop rapidly following antigenic exposure, making the shortened window critical for preventing delayed hemolytic reactions 4

Extended Antigen Typing Recommendations

When to Perform Extended Typing

Beyond basic ABO/RhD, perform extended red cell antigen profiling for Rh antigens (C, c, E, e) and K antigen in the following situations:

• All patients with sickle cell disease requiring transfusion (strong recommendation) 5
• Patients requiring chronic transfusion therapy 6
• Patients with positive antibody screens to expedite antibody identification 5
• Women of childbearing potential to assess risk for hemolytic disease of the fetus and newborn 6

Rationale: Antibodies to C, E, and K antigens are the most common specificities complicating transfusion, and extended matching reduces alloimmunization from 3.1 per 100 units to 0.9 per 100 units 6

Methodology Selection

DNA-based genotyping is preferred over serologic phenotyping when:

• Recent transfusion interferes with serologic testing 5
• Allo- or autoantibodies interfere with interpretation 5
• Greater accuracy is needed for C and Fy(b) antigen matching 5
• The patient has complex antibody patterns 5

However, acknowledge that genotyping is often a referral test with longer turnaround times, so serologic testing at first encounter remains practical for most hospital transfusion services 5

Emergency Situations with Unknown Blood Type

Blood Selection Algorithm

For emergency transfusion when blood type is unknown:

• Women of childbearing potential: Use Group O RhD-negative red blood cells exclusively to prevent RhD alloimmunization and future hemolytic disease of the fetus and newborn 7
• Males, postmenopausal females, and adults without childbearing potential: Group O RhD-positive blood is acceptable and preferred to conserve scarce RhD-negative inventory 7

Transition Strategy:

• Switch to group-specific blood within 10-15 minutes once laboratory receives a properly labeled sample 7
• Blood grouping can be performed in approximately 10 minutes 7
• In massive bleeding, patients have minimal circulating antibodies and usually accept group-specific blood without reaction 7

Rapid Testing Options

A rapid ABO and RhD test (such as Eldoncard) demonstrates 99.5% concordance with reference laboratory testing, with 99.3% sensitivity and 98.2% negative predictive value for RhD typing 8. This can be used in emergency departments to guide blood product selection during resuscitation 8.

Reporting Standards

Essential Elements to Report

Every blood grouping report must include:

• ABO group (A, B, AB, or O) confirmed by both forward and reverse typing 1
• RhD type (positive or negative) 1
• Antibody screen result (positive or negative) 1, 3
• Date and time of sample collection 4
• Sample validity period based on transfusion/pregnancy history 4

For positive antibody screens:

• Identify specific antibody specificities (e.g., anti-K, anti-E, anti-c) 3
• The most frequently identified alloantibodies are anti-K (32%) and anti-E (30.8%) 3

For extended phenotyping when performed:

• Report Rh antigens: C, c, E, e 5
• Report K antigen status 5
• Consider reporting Jk(a)/Jk(b), Fy(a)/Fy(b), M/N, S/s when indicated 5

Critical Safety Considerations

Common Pitfalls to Avoid:

• Most transfusion-related morbidity results from incorrect blood administration and patient identification errors, not blood group incompatibility 7
• Always use four core patient identifiers 7
• RhD variants may produce discrepant results between rapid tests and reference laboratory methods; when serological characteristics suggest a variant, perform genotyping 8
• Never rely on historical samples alone for recently transfused or pregnant patients—the 72-hour rule is mandatory 4
• Patients with GATA mutation in the ACKR1 gene are not at risk for anti-Fy(b) and do not require Fy(b)-negative red cells 5

Special Population Alerts:

• Multitransfused patients have the highest alloimmunization rates and can develop new antibodies rapidly 4
• Patients with hybrid RHDDIIIa-CE(4-7)-D or RHCECeRN alleles encoding partial C antigen should receive C-negative red cells 5