[Establishment and Applications of Double-Fluorescent Protein Allo-Transplantation Mice Model]

To establish allo-transplantation model by using mRFP⁺ to eGFP⁺ transgenic mice and to observe the distribution of donor cells and donor-recipient cellular interaction in the bone marrow after semi-solid decalcification (SSD).

After myeloablative irradiation, C57BL/6 female eGFP⁺ transgenic mice were infused with (5 × 10⁶) bone marrow cells from FVB male donor mice through tail vein. The control group was infused with PBS. Then the general conditions, engraftment level, hematopoietic recovery, incidence of GVHD and survival of recipients were evaluated after transplantation. In the recovery process, SSD was used to treat the femora before observing the cells distribution, morphology and interaction by confocal microscopy directly or after making frozen section.

WBC of recipient eGFP⁺ mice was recovered on (20 ± 3.07) d, (93.94 ± 1.59)% in peripheral cells were RFP⁺ cells (n = 10), GVHD happened in 4 of 10 mice within 1 month. During SSD, the hard components were replaced gradually and RFP⁺ cells could be seen mainly in the bone trabecula and surrounded by eGFP⁺ cells under confocal microscope, their interactions could be further observed clearly in bone marrow microenvironment in three-dimensional reconstruction.

The double fluorescent allo-transplantation mouse model successfully established, by means of our novel protocol named SSD, the donor and recipient cell location and their interaction can be visually observed, which provides the basis for clinical studies on the distribution and homing of donor cells, and some related explorations after transplantation.