Testing

This is arguably one of the most critical parts of ensuring blood safety, acting as a multi-layered shield to protect recipients. After blood is collected, but before it can be labeled and released for transfusion, samples from every single donation undergo a battery of tests mandated by the FDA and upheld by AABB standards

Think of testing in two main categories:

Infectious Disease Marker Testing: To prevent transmitting diseases
Immunohematology Testing: To ensure blood type compatibility

Infectious Disease Marker Testing (Transfusion-Transmitted Infections - TTIs)

Goal: To detect the presence of specific viruses or bacteria in the donated blood that could harm a recipient
Mandate: Required by FDA (21 CFR 610 Subpart E) for every donation. AABB standards mirror these requirements
Methodology: Uses FDA-licensed screening test kits. Two main technologies are employed:
- Serological Tests: Detect either antibodies produced by the donor’s immune system in response to an infection, or antigens, which are proteins directly from the infectious agent itself (like Hepatitis B surface antigen - HBsAg)
- Nucleic Acid Testing (NAT): A highly sensitive molecular method that detects the genetic material (RNA or DNA) of the virus. NAT is crucial because it can detect infections much earlier than antibody tests, during the “window period” when someone is infected but hasn’t yet produced detectable antibodies. This significantly enhances safety. NAT is often performed on pooled samples first; if a pool tests positive, individual units within the pool are then tested
Required Tests (Current US FDA Requirements)
- HIV (Human Immunodeficiency Virus)
  - HIV-1/2 Antibody (detects antibodies to both major types)
  - HIV-1 NAT (detects viral RNA)
- HCV (Hepatitis C Virus)
  - HCV Antibody (detects antibodies)
  - HCV NAT (detects viral RNA)
- HBV (Hepatitis B Virus)
  - HBsAg (Hepatitis B Surface Antigen - detects part of the virus itself)
  - Anti-HBc (Antibody to Hepatitis B Core antigen - indicates current or past infection)
  - HBV NAT (detects viral DNA)
- HTLV (Human T-lymphotropic Virus)
  - HTLV-I/II Antibody (detects antibodies)
- Syphilis
  - Serological test (detects antibodies to Treponema pallidum)
- WNV (West Nile Virus)
  - WNV NAT (detects viral RNA; crucial as WNV often causes asymptomatic infection)
- Zika Virus
  - Zika Virus NAT (detects viral RNA; requirement often based on FDA guidance and risk)
- Trypanosoma cruzi (Chagas Disease)
  - Antibody test (required at least once per donor; detects antibodies to the parasite)
- Babesia microti:
  - NAT or Antibody test (required in states designated by FDA as endemic, recommended elsewhere; detects the parasite common in tick-borne infections)
Interpretation & Action
- If any screening test is reactive (positive), the unit of blood is quarantined and discarded. It cannot be used for transfusion
- Confirmatory testing is usually performed on reactive units to verify the result
- The donor is notified of confirmed positive results and placed on the appropriate deferral list (often permanent for HIV, HCV, HBV)
- “Lookback” procedures may be initiated to identify recipients of previous donations from that donor if applicable

Immunohematology Testing (Blood Group Serology)

Goal: To determine the donor’s blood type (ABO and Rh) and screen for unexpected red cell antibodies to ensure compatibility with potential recipients and prevent hemolytic transfusion reactions
Mandate: Required by FDA (21 CFR 640) and AABB standards
Required Tests
- ABO Grouping
  - Forward Type: Tests the donor’s red cells with known anti-A and anti-B reagents to detect A and/or B antigens
  - Reverse Type: Tests the donor’s plasma/serum with known A1 and B red cells to detect anti-A and/or anti-B antibodies
  - Crucial Step: The forward and reverse type results must agree for the ABO group to be validly assigned. Any discrepancy requires investigation
- Rh Typing
  - Tests the donor’s red cells with anti-D reagent to determine if the D antigen (the primary Rh antigen) is present (Rh positive) or absent (Rh negative)
  - Weak D Testing: Required by AABB standards (and implicitly by FDA outcomes) for donors who initially test Rh negative with standard anti-D. If the weak D test is positive, the unit is labeled as Rh Positive. If negative, it’s labeled Rh Negative. This prevents Rh-negative recipients from being exposed to the D antigen
Additional Immunohematology Testing
- Antibody Screen (Detection of Unexpected Red Cell Antibodies)
  - Tests the donor’s plasma/serum against screening cells (group O cells with known antigen profiles) to detect clinically significant IgG antibodies (other than the expected anti-A or anti-B)
  - Required by AABB for donors with a history of pregnancy or transfusion. Many centers perform it on all donors
  - Why?: If unexpected antibodies are present:
    - Plasma-containing components (FFP, Platelets) should ideally not be given to recipients whose red cells have the corresponding antigen
    - Red Blood Cell units are generally safe (as most plasma is removed), but the information is valuable. The unit must be labeled as containing the antibody
    - These donors may be recruited for reagent red cell manufacturing
- Red Cell Antigen Phenotyping
  - Not routinely required for all donors, but may be performed to identify donors negative for common antigens (like Kell, Duffy, Kidd, S, s) to build an inventory for patients with specific antibodies
Interpretation & Action
- ABO and Rh type results are critical for correct unit labeling
- Positive antibody screen requires antibody identification. Units are labeled accordingly
- Historical records for repeat donors are checked to ensure consistency in ABO/Rh typing

Quality Control Testing of Components

While not donor testing per se, it’s part of processing. Blood centers perform QC tests on a subset of components after preparation to ensure they meet standards (e.g., platelet count and pH in platelets, residual WBCs in leukoreduced units, hematocrit in RBCs)

Key Takeaway

Testing is a non-negotiable, highly regulated part of blood processing. It combines sensitive infectious disease screening (especially using NAT) with precise immunohematology testing. This rigorous process is essential for ensuring that the blood components released from the blood center are as safe as possible and compatible for the intended recipients. No unit gets labeled and released until all required tests are completed and acceptable

Key Terms

Transfusion-Transmitted Infection (TTI): An infection acquired through a blood transfusion
Serological Testing: Testing blood serum/plasma for the presence of antibodies or antigens
Nucleic Acid Testing (NAT): Molecular testing that detects the DNA or RNA of infectious agents like viruses
Window Period: The time between infection and when a test (usually antibody-based) can detect evidence of that infection. NAT significantly shortens this period
Antigen: A substance (often a protein) on the surface of a cell or pathogen that can trigger an immune response
Antibody: A protein produced by the immune system in response to an antigen
ABO Grouping: Determining blood type based on the presence or absence of A and B antigens on red cells and anti-A and anti-B antibodies in plasma
Rh Typing: Determining the presence or absence of the D antigen on red cells
Weak D: Red cells that express the D antigen weakly, requiring special testing methods for detection. Donors positive for weak D are considered Rh Positive
Antibody Screen: A test to detect unexpected clinically significant red cell antibodies in plasma/serum
Lookback: The process of identifying recipients of blood from a donor who later tested positive for a relevant TTI, to notify and test the recipients
Quarantine: Holding blood units in a designated area, unavailable for issue, until all required testing and processing steps are completed and reviewed

FDA, AABB Requirements

Labeling