Figure 4
Human PBMC label efficiently with a 19F-PFC cell-labeling agent under GMP-compliant conditions and transported without significant loss of viability. (a) Following overnight culturing of PBMC with and without the 19F-PFC cell-labeling agent (5 mg/mL) under GMP-compliant conditions, labeled and unlabeled PBMC were transported back to London, Canada. The viability of 19F-PFC-labeled PBMC and unlabeled PBMC was determined using 7-AAD staining before and after transport. There was no significant decrease in viability because of transport (approximately 2.5 hours for a 250 km distance) as well as no significant difference in viability between labeled and unlabeled cells. (b) PBMC were labeled overnight with the red fluorescent version of the 19F-PFC cell-labeling agent. Half of the 19F-PFC labeled cells were stained for CD45 and for viability and analyzed by flow cytometry which revealed that 99.8% of PBMC were positively labeled with the red fluorescent version of the 19F-PFC agent compared to unlabeled PBMC from the same donor (blue histogram). (c) The second of half of the labeled PBMC were processed and stained for CD45 and viability ~3 hours later followed by flow cytometry which revealed that 99% of the PBMC retained the red fluorescent 19F-PFC compared to unlabeled PBMC from the same donor (blue histogram). (d) In order to quantify in vivo cell migration, a quantitative assessment of 19F-PFC incorporation must also be ascertained using NMR spectroscopy on a known number of PBMC. For the 5 participants analyzed, an average of 6.17 × 1010 19F spins was incorporated per cell. For the last three participants, an average of 7.55 × 1010 19F spins were incorporated per cell and are denoted by the white circles and are the participants for which MR image data is provided.
