Ziel/Aim:
To evaluate in vivo dual PET/FACS mAb-labeling of neutrophils for cell tracking in small rodents without the need for adoptive cell transfer.
Methodik/Methods:
To investigate neutrophil migration, Cu-64-NODAGA-AF700 – 1A8, a dual-labeled monoclonal antibody (mAb) for in vivo PET and ex vivo FACS (AlexaFluor700, AF700) was used (20 µg, 10MBq). Static PET/MR measurements were conducted in 20-months-old APPPS1 and C57BL/6 (WT) mice at 3h, 24h and 48h post injection (pi, n = 5). Neutrophil migration was stimulated 24h pi with G-CSF. At 48h pi, FACS analysis was performed. For control experiments, neutrophils were depleted with 100 µg/kg 1A8 in APPPS1 and WT mice (n = 4). Two-tailed Student's t-test was used, p < 0.05 was considered statistically significant.
Ergebnisse/Results:
PET analysis revealed significantly elevated uptake 48h pi in brain (WT: 0.61 ± 0.07%ID/cc; APPPS1: 0.75 ± 0.06%ID/cc, n = 5; p = 0.01) and spleen of APPPS1 mice (WT: 2.22 ± 0.53%ID/cc; APPPS1: 3.79 ± 0.73%ID/cc; p = 0.004). FACS analysis did not confirm PET results and showed no significant difference in Ly6G+AF700+ populations in APPPS1 compared to WT brains (WT: 5.62 ± 8.54% of CD45.2; APPPS1: 1.95 ± 1.39% of CD45.2; p = 0.37). In spleen and bone marrow, no significant differences in AF700+ populations were observed. Control PET experiments showed markedly reduced uptake in spleen and bone marrow, but not in brain compared to non-depleted condition. Similar results were detectable for Ly6G+AF700+ populations in spleen and bone marrow; however, values were significantly higher in APPPS1 brains (WT: 0.20 ± 0.08% of CD45.2; APPPS1: 1.23 ± 0.56% of CD45.2; p = 0.01).
Schlussfolgerungen/Conclusions:
Although PET and FACS revealed dissimilar results in brain analysis and further evaluations are needed, we established a promising technique to dynamically investigate in vivo cell migration – and cross-correlate it with ex vivo FACS – to the CNS, but potentially also to other organs.
