What is the role of the indirect agglutination test (IAT) in pre-transfusion testing?

Indirect Agglutination Test (IAT) in Pre-Transfusion Testing

Essential Role as the Gold Standard

The indirect antiglobulin test (IAT) is the cornerstone of pre-transfusion antibody screening and must be performed on all patients requiring transfusion to detect clinically significant red blood cell antibodies that can cause hemolytic transfusion reactions. 1

The IAT serves three critical functions in pre-transfusion testing:

• Antibody screening: Detects unexpected alloantibodies in patient plasma that could react with donor red cells 1
• Antibody identification: Determines the specificity of detected antibodies to guide compatible blood selection 2
• Crossmatching: Confirms compatibility between patient plasma and specific donor units before transfusion 2

Technical Methodology and Enhancement

Standard IAT Protocol

The IAT detects IgG antibodies and complement components bound to red blood cells after incubation at 37°C 3. The test requires:

• Incubation of patient serum with reagent red cells at 37°C 3
• Washing to remove unbound antibodies 4
• Addition of anti-human globulin (AHG) reagent to detect bound antibodies 4
• Reading for agglutination indicating antibody presence 4

Potentiation Methods

Polyethylene glycol-enhanced IAT (PEG-IAT) is superior to albumin-enhanced IAT (Alb-IAT) for detecting clinically significant antibodies while reducing false-positive reactions. 4

• PEG-IAT more frequently detects critical antibodies including anti-E, anti-Fy(b), and anti-Jk(a) 4
• PEG-IAT reduces detection of clinically insignificant antibodies such as anti-Le(b) and anti-P1 4
• PEG-IAT decreases the incidence of delayed hemolytic transfusion reactions (0.12% vs 0.30% with Alb-IAT) 4

Modern Gel Technology

Column agglutination technology (gel test) is sufficiently sensitive that 37°C saline testing can be safely omitted from the pre-transfusion protocol. 3

• All antibodies detected by 37°C saline testing are also detected by gel-based IAT 3
• Gel methods provide standardized, objective reading of results 3
• The gel system eliminates the need for redundant testing phases 3

Critical Timing Requirements

Group-specific blood should be available within 15-20 minutes of the laboratory receiving a properly labeled sample, while complete antibody screening and crossmatching typically requires 45-60 minutes. 5, 2

For life-threatening hemorrhage:

• Group O RhD negative red cells should be immediately available without IAT testing 2, 5
• Group O RhD positive is acceptable for males and postmenopausal females to conserve RhD negative inventory 5
• Transition to group-specific blood as soon as basic ABO/RhD typing is complete (10-15 minutes) 5

Special Populations Requiring Extended Testing

Sickle Cell Disease Patients

Patients with sickle cell disease require extended red cell antigen phenotyping beyond standard ABO/RhD typing to prevent alloimmunization. 2

• Extended phenotyping should include Rh (C/c, E/e), K, Jk(a)/Jk(b), Fy(a)/Fy(b), M/N, and S/s antigens 2
• This extended profile expedites antibody identification when patients develop positive antibody screens 2
• Red cell genotyping provides more comprehensive information than serologic methods and improves accuracy 2
• The extended phenotype facilitates finding compatible blood for patients with multiple alloantibodies 2

Resolving Complex IAT Problems

Panagglutination

When patient serum reacts with all screening and panel cells (panagglutination), a systematic approach is required 6:

• First assess: Intensity of reactivity and whether autocontrol is positive or negative 6
• Determine cause: Distinguish between autoantibody, single alloantibody, multiple alloantibodies, or antibody to high-incidence antigen 6
• Critical question: Identify any clinically significant alloantibodies masked by the panagglutination 6

Drug Interference

Anti-CD38 monoclonal antibodies (used for multiple myeloma treatment) cause agglutination in IAT by binding to red blood cells, requiring specialized techniques to resolve the interference. 7

• The drug interference causes delays in issuing compatible blood to anemic patients 7
• Multiple methods exist to eliminate or bypass anti-CD38 effects, but no universal solution exists 7
• Each testing site must tailor an approach based on their specific capabilities and patient population 7

Techniques to Avoid

Two-stage papain-IAT and LISS direct agglutination should NOT be used for routine pre-transfusion antibody screening. 8

• These methods detect predominantly clinically insignificant antibodies (anti-Lewis, anti-P1) 8
• They produce 81% of unwanted positive reactivities that delay transfusion 8
• The additional antibodies detected (mostly anti-Lewis) rarely have clinical significance 8

Documentation and Safety

All IAT results must be documented in the patient record with full traceability 2, 1:

• Document the indication for testing, methods used, and results obtained 1
• Maintain 100% traceability of all blood components as a legal requirement 2
• Report any adverse reactions according to local protocols and national hemovigilance systems 1
• Inform patients they have received blood components before discharge, as this removes them from the donor pool 2