CHT Crossmatch Procedures & Interpretation 1 — Questions and Answers
Question 1: What is the primary purpose of performing a pretransplant crossmatch?
- To detect preformed donor-specific HLA antibodies in the recipient's serum (Correct answer)
- To determine the donor's ABO blood type compatibility
- To assess the recipient's renal function before transplant
- To evaluate donor-recipient body size compatibility
Correct answer: To detect preformed donor-specific HLA antibodies in the recipient's serum
The pretransplant crossmatch detects preformed donor-specific antibodies (DSAs) in the recipient that could cause hyperacute rejection upon transplantation.
Question 2: In a complement-dependent cytotoxicity (CDC) crossmatch, cell death is assessed by:
- Flow cytometry using fluorescent cell-surface markers
- Exclusion of a vital dye such as eosin Y or trypan blue (Correct answer)
- ELISA optical density readings at 450 nm
- Luminex mean fluorescence intensity (MFI) values
Correct answer: Exclusion of a vital dye such as eosin Y or trypan blue
CDC crossmatch uses vital dye exclusion — dead cells with compromised membranes take up the dye, allowing percent cytotoxicity to be calculated.
Question 3: A positive T-cell crossmatch using current recipient serum is clinically significant because it indicates:
- ABO blood group incompatibility between donor and recipient
- Donor-specific anti-HLA class I antibodies in the recipient (Correct answer)
- The presence of non-HLA autoantibodies in the recipient
- Recipient sensitization to HLA class II antigens exclusively
Correct answer: Donor-specific anti-HLA class I antibodies in the recipient
T cells express HLA class I antigens (A, B, C), so a positive T-cell crossmatch indicates the recipient has preformed antibodies against donor HLA class I antigens.
Question 4: Dithiothreitol (DTT) treatment of recipient serum before crossmatch testing is used to:
- Enhance complement pathway activation in the assay
- Inactivate IgM antibodies by reducing their disulfide bonds (Correct answer)
- Remove HLA class II antigens from the test mixture
- Increase lymphocyte viability during the incubation period
Correct answer: Inactivate IgM antibodies by reducing their disulfide bonds
DTT cleaves the disulfide bonds that hold the pentameric IgM structure together, selectively inactivating IgM and allowing identification of IgG-mediated crossmatch reactivity.
Question 5: Which best describes an auto-crossmatch and its clinical use?
- Testing recipient serum against cells from a random unrelated donor
- Testing recipient serum against the recipient's own lymphocytes (Correct answer)
- Testing donor serum against recipient lymphocytes in reverse
- Testing archived historical serum against current donor cells
Correct answer: Testing recipient serum against the recipient's own lymphocytes
An auto-crossmatch tests recipient serum against the recipient's own cells to detect autoantibodies, which can cause false-positive crossmatch results if not identified.
Question 6: The Amos-modified CDC crossmatch improves specificity over the standard CDC by:
- Adding extra rabbit complement to increase lytic activity
- Including wash steps to remove nonspecific serum components before complement addition (Correct answer)
- Substituting human complement for rabbit complement
- Increasing the serum-to-cell ratio in the incubation step
Correct answer: Including wash steps to remove nonspecific serum components before complement addition
Washing steps in the Amos-modified CDC remove unbound antibodies and serum proteins that may activate complement nonspecifically, reducing false-positive results.
Question 7: Pronase treatment of donor lymphocytes before CDC crossmatch testing enhances sensitivity by:
- Removing HLA class I antigens to reduce background reactivity
- Stripping B-cell surface markers such as CD19 and CD20
- Cleaving complement regulatory proteins (CD55, CD59) from the cell surface (Correct answer)
- Inactivating residual IgG antibody present in the serum
Correct answer: Cleaving complement regulatory proteins (CD55, CD59) from the cell surface
Pronase is a protease that removes complement inhibitory proteins (CD55/DAF and CD59/protectin) from the cell surface, making cells more susceptible to complement-mediated lysis.
CHT Crossmatch Procedures & Interpretation 1
What is the primary purpose of performing a pretransplant crossmatch?