Rapid spectrophotometric technique for quantifying iron in cells labeled with superparamagnetic iron oxide nanoparticles: potential translation to the clinic

Abstract

Labeling cells with superparamagnetic iron oxide (SPIO) nanoparticles provides the ability to track cells by Magnetic Resonance Imaging. Quantifying intracellular iron concentration in SPIO labeled cells would allow for the comparison of agents and techniques used to magnetically label cells. Here we describe a rapid spectrophotometric technique (ST) to quantify iron content of SPIO labeled cells, circumventing the previous requirement of an overnight acid digestion. Following lysis with 10% SDS of magnetically labeled cells, quantification of SPIO doped or labeled cells was performed using commonly available spectrophotometric instrument(s) by comparing absorptions at 370 and 750 nm with correction for turbidity of cellular products to determine iron content of each sample. Standard curves demonstrated high linear correlation (R2 = 0.998) between absorbance spectra of iron oxide nanoparticles and concentration in known SPIO doped cells. Comparisons of the ST to ICP-MS or NMR relaxometric (R2) determinations of intracellular iron contents in SPIO containing samples resulted in significant linear correlation between the techniques (R2 vs. ST, R2>0.992, p<0.0001, ST vs. ICP-MS, R2>0.995, p<0.0001) with the limit of detection of ST for iron = 0.66μg/ml. We have developed a rapid straightforward protocol that does not require overnight acid digestion for quantifying iron oxide content in magnetically labeled cells using readily available analytic instrumentation that should greatly expedite advances in comparing SPIO agents and protocols for labeling cells.

Physicochemical characterization of ferumoxytol, heparin and protamine nanocomplexes for improved magnetic labeling of stem cells

“…Intracellular iron content was obtained by a rapid spectroscopic technique that following digestion of 10 6 /ml labeled or control cells with sodium dodecyl sulfate (SDS 10% Manufacturer) for 5min and the difference in light absorbed at 370 and 750nm using UV/Vis spectrometer (PerkinElmer Lambda 25, Shelton, CT, USA) are compared to standard curve to determine iron content as previously described[22]. Iron contents were performed in triplicate and expressed as mean and standard deviation.…”

Flow cytometry for intracellular SPION quantification: specificity and sensitivity in&nbsp;comparison with spectroscopic methods

“…22 In detail, 50 μL of cell lysate (2×10 5 cells) and SPION standards were pipetted into 96-well plates (Techno Plastic Products AG, Trasadingen, Switzerland) and the OD 370 was measured in a spectrophotometer (FilterMax F5, Molecular Devices, Sunnyvale, CA, USA). Standard dilutions of SPIONs enabled an absolute quantification of the iron nanoparticle concentration in the cell lysates.…”

“…To quantify the cellular SPION content, we modified a photometric technique recently published by Dadashzadeh et al 22 Using this method, the absorption maximum for SPIONs should be observable at 370 nm. To ensure that this also applies to the SPIONs used in this study, we prepared SPION dilutions of 30 μg Fe /mL in 10% sodium dodecyl sulfate and monitored the corresponding absorption spectra between 200 nm and 900 nm with a stepwise increase of 2 nm.…”

“…22 As the coating of SPIONs might influence their absorption behavior, we investigated the reproducibility of this method for SEON nanoparticles coated with either lauric acid or lauric acid and bovine serum albumin in parallel with Rienso ® particles coated with PSC. We observed similar absorption spectra and an absorption maximum at 370 nm for all three SPION types, indicating only a very slight influence of the coating on absorption intensity at OD 370 ( Figure S2A).…”

Electron paramagnetic resonance: a powerful tool to support magnetic resonance imaging research

“…To achieve that goal, radioactive 59 Fe-SPIO can be used, but this requires the chemical synthesis of radiolabeled nanoparticles (139) which is cumbersome. Inductively coupled plasma mass spectrometry (ICP-MS) or inductively coupled plasma optical emission spectrometry (ICP-OES) are alternatives that provide a sensitive quantification of elemental iron in tissues (150)(151)(152). However, it should be noted that this method is unable to discriminate endogenous iron from exogenous iron oxide nanoparticles.…”