Signal and Measurement Considerations for Human Translation of Diffuse in vivo Flow Cytometry

Abstract

SignificanceDiffuse in vivo flow cytometry (DiFC) is an emerging technology for fluorescence detection of rare circulating cells directly in large deep-seated blood vessels in mice. Because DiFC uses highly scattered light, in principle it could be translated to human use. However, an open question is whether fluorescent signals from single cells would be detectable in human-scale anatomies.AimSuitable blood vessels in a human wrist or forearm are at a depth of approximately 2-4 mm. The aim of this work was to study the impact of DiFC instrument geometry and wavelength on the detected DiFC signal and on the maximum depth of detection of a moving cell.ApproachWe used Monte Carlo simulations to compute Jacobian (sensitivity) matrices for a range of source-detector separations and tissue optical properties over the visible and near infrared (NIR) spectrum. We performed experimental measurements with three available versions of DiFC (488 nm, 640 nm, and 780 nm), fluorescent microspheres, and tissue mimicking optical flow phantoms. We used both computational and experimental data to estimate the maximum depth of detection at each combination of settings.Results and ConclusionsFor the DiFC detection problem, our analysis showed that for deep-seated blood vessels, the maximum sensitivity was obtained with NIR light (780 nm) and 3 mm source-and-detector separation. These results suggest that - in combination with a suitable molecularly targeted fluorescent probes - circulating cells and nanosensors could in principle be detectable in circulation in humans.

Near Infrared Diffuse In Vivo Flow Cytometry

“…In addition to the advantages of NIR-DiFC described above, as summarized in fig. 4 we observed that tissue autofluorescence is markedly reduced and the signal is free of motion artifacts (35) compared to our pre-clinical GFP-compatible b-DiFC instrument (31). Use of blue laser light is widely understood to yield higher biological tissue autofluorescence than near-infrared light (49).…”

“…We first tested NIR-DiFC using an optical flow phantom ( Fig. 2a ), which we have shown previously approximates the optical properties of bulk biological tissue in the NIR range (35). Peaks were never observed in control solutions of either PBS or unlabeled cells; a representative NIR-DiFC data scan for unlabeled L1210A cells is shown in fig 2b .…”

“…The rationale was three-fold. First, NIR-DiFC should allow detection of CTCs in blood vessels 2-4 mm deep using our DiFC approach (35). Hence, major blood vessels in the wrist or forearm of a human should be accessible with NIR-DiFC (34).…”

“…We first tested NIR-DiFC using an optical flow phantom (Fig. 2a), which we have shown previously approximates the optical properties of bulk biological tissue in the NIR range (35).…”

“…NIR light is known to experience lower attenuation (scattering and absorption) in biological tissue than visible light, and therefore is expected to be better suited to larger anatomies (33). We recently showed that NIR light would enable detection of well-labeled CTCs in large blood vessels 2-4 mm below the surface, which would be suitable for measurement of cells in the human wrist or forearm (34, 35). Hence, development of NIR-DiFC is a logical first step in potential human translation of the technique.…”