Figure 6, highlighting the changes in contrast for a given molar concentration. Likewise, T * relaxivity was similar for all samples, with no specific trend related to 2 core size (Figure 6b). The T * relaxivity observed in cells, however, was higher than 2 that obtained in solution, particularly for cells labelled with SPIONs with a core size of 6.7 and 8.4 nm (Tables S3 & S4). This is probably because once internalised in size,3 cells, SPIONs are often packed in vesicles well over 500 nm in thus leading to correspondingly large magnetic field inhomogeneities. This can be seen in the images shown in Figure 6b, where negative contrast is often punctuated, rather than uniform as seen for colloidally suspended particles. Importantly, this clustering of the SPIONs seems to dwarf any effect related to particle size, highlighting that obtaining high intracellular concentrations of iron in a safe manner is likely to be more important than obtaining particles of extremely high relaxivity.

## Conclusions

We have shown that the amount of polymer used in a co-precipitation reaction can impact not only the physical properties of SPIONs, but also their efficiency as stem cell tracking agents for MRI. Decreasing the ratio of FITC-DEAE-dextran to iron salts used in the synthesis yielded SPIONs with larger core size and less polymer in the outer shell, leading to a significant impact on the uptake of SPIONs into MSCs. At a mL-1, nominal labelling concentration of 50 µg nearly a five-fold increase in uptake could be achieved for SPIONs with mean core sizes of 9.9 nm and ~ 30 % polymer content compared to those SPIONs with a mean core size of 6.7 nm and 73 % polymer content, without altering cell viability. This suggests that core size and overall polymer content can largely affect uptake, as well as surface charge. The increase in uptake was reflected in the amount of negative contrast observed by MRI. We also demonstrated how the physicochemical properties of SPIONs can change significantly once exposed to physiological medium or internalised into MSCs. The superior uptake and contrast generated with particles synthesised with the conditions presented here suggest these SPIONs are strong candidates for in vivo imaging in clinical and pre-clinical settings, warranting further studies involving a broader spectrum of cell lines and in vivo models.

Experimental Characterisation Powder X-Ray diffraction (pXRD). Measurements were carried out on a Panalytical Powder X-Ray diffraction (pXRD). Measurements were carried out on a Panalytical X’pert Pro multipurpose diffractometer with a Co Kα source (λ = 1.78 Å). Patterns were measured over 20-120° 2θ for 2 hours (step size 0.033°, time per step 295.3 s and scan speed 0.014 °/s). The mean core diameter (D ) was calculated by using the fit m profile tool (pseudo Voigt peak profile) on X’Pert Highscore plus software to obtain β, the full width half maximum (FWHM) of the 311 Bragg reflection and applying the Scherrer equation:

where where K = unknown shape factor and θ = the Bragg angle. For the stacked B and overlapped figures, the background was subtracted for clarity. A Pawley refinement was performed using Topas academic software to confirm the purity of the crystalline phase. The lattice parameter values were taken from the known structure of