Advances in Fluorescence‐BasedIn vivoFlow Cytometry for Cancer Applications

Circulating Tumor Cell Detection and Capture by Photoacoustic Flow Cytometry in Vivo and ex Vivo

“…, as in conventional FC) [ 38 , 39 , 40 , 41 , 42 , 46 , 47 , 48 , 49 ] and photothermal (PT) and photoacoustic (PA) detection methods [ 28 , 33 , 34 , 35 , 36 , 37 ], and, recently the combination of these methods [ 28 , 29 , 49 ]. A brief history and the features and challenges of this next generation of FC have been described in reviews and book chapters ([ 33 , 34 , 43 , 44 , 45 , 46 , 47 , 48 ] and references there) with a focus on early work in this field and broad applications, including circulating RBCs and WBCs in different functional states (e.g., normal, apoptotic), infections ( E. coli and S. aureus ), sickle cells, blood rheology, and pharmacokinetics of nanoparticles (NPs), drug carriers, and dyes and other contrast agents. Here, we summarize recent advances of in vivo FC platform with a focus on detecting CTCs with PAFC methods.…”

“…In vivo FC, with fluorescence detection of CTCs, can be built on the basis of conventional and confocal microscope schematics, single-, two-, or multiphoton excitation, and standard fluorescent labels, as in conventional FC in vitro [ 38 , 39 , 46 ]. In the confocal scheme, fluorescent signals from the cell population of interest are recorded as the cells pass through a slit of a continuous-wave (CW) laser (e.g., He-Ne) light focused across 20–50 µm mouse ear blood vessels.…”

“…Recent advances include the use of two-color schematics and fiber-based laser delivery [ 42 , 47 ]. In vivo fluorescence flow cytometry (FFC) was successfully used for in vivo detection of labeled CTCs in animal models [ 46 , 47 , 48 , 49 ]. Nevertheless, the use of fluorescent labeling raises concerns due to the cytotoxicity of fluorescent tags and their usefulness for assessing only superficial microvessels (≤50–100 µm) with slow flow velocities (1–5 mm/s) due to the influence of a strong autofluorescent background.…”

“…In addition, invasive extraction of blood from a living organism may alter CTC parameters (e.g., signaling, epigenetic states, metabolic activities, or morphology) and prevent the long-term study of CTC properties (e.g., CTC-blood cell interactions, aggregation, rolling, or adhesion) in their natural biological environment. These problems can be solved by assessing a significantly larger volume of blood up to a patient’s entire blood volume with in vivo FC, whose principles were proposed in 2004 simultaneously by Zharov and other groups [ 33 , 34 , 35 , 36 , 37 , 38 , 39 , 40 , 41 , 42 , 43 , 44 , 45 , 46 , 47 , 48 , 49 ].…”

In vivo flow cytometry: A horizon of opportunities

“…PT and PA methods in combination with high speed optical imaging and Raman spectroscopy were also applied by Zharov's team on quantitative monitoring of individual cells in lymph flow (19,22,26,28). The advances in in vivo FCM were also made by other researchers and research groups (52, 58–79), including Irene Georgakoudi (Tufts University), Theodore Norris (University of Michigan), Xunbin Wei (Fudan University), Eric Tkaczyk (University of Tartu), and Anja Hauser (Deutsches Rheumaforschungszentrum). These advances were made mostly in the fluorescent‐based in vivo FCM (58, 61–64), confocal fluorescence (59, 60, 62) and backscattering (65, 66) microscopy, multiphoton laser scanning microscopy (63, 67–78), and coherent anti‐Stokes Raman scattering (CARS) (79) techniques.…”