Various techniques are available for distinguishing donor from host cells evaluating the efficacy of conditioning regimen for experimental bone marrow transplantation (BMT). Techniques include the use of extracellular immunological markers, such as Ly5 (CD45), and intracellular biochemical markers, such as glucose-phosphate-isomerase (Gpi). Because Ly5 is an extracellular protein, the disparity between donor (Ly5.1) and host (Ly5.2) antigens may induce a weak immune response whereas with Gpi, no immune response is expected. This difference may be of particular concern in experimental transplantation approaches that use minimal conditioning such as low-dose total body irradiation (TBI). Such mild conditioning may not induce the immunosuppression required to overcome host rejection of Ly5 disparate cells.

To compare the relative engraftment of Ly5.1 and Gpi-1^a donor marrow, B6 (Gpi-1^b/Ly5.2) mice were irradiated with low-level TBI (0-6 Gy) and transplanted with several bone marrow (BM) doses (2 × 10⁶-5 × 10⁷ cells). At 8, 26, and 52 weeks post-BMT, the level of donor engraftment was measured using flow cytometry (Ly5) or Gpi-electrophoresis.

Lower engraftment levels were found in mice transplanted with Ly5 congenic BM in groups given low-dose TBI (≤4 Gy) and/or low doses of BM cells (BMC) (2 × 10⁶). However, when higher TBI or BMC doses were used, similar engraftment levels were found, suggesting sufficient immune suppression to allow equal engraftment of both sources of BM.

These data suggest that even a minor phenotypic disparity between donor and host, such as Ly5, may necessitate high-dose TBI to prevent rejection. The combination of low-dose TBI or other nonmyeloablative conditioning strategies with small numbers of BMC may lead to reduced engraftment when extracellular immunological markers such as Ly5 are used for transplantation studies. Therefore, small immunological differences must be considered when using the Ly5 marker for engraftment.