In this article, we demonstrate the potential of a microfluidic chip for the differentiation of immunologically stained blood cells. To this end, white blood cells stained with antibodies typically applied for the determination of the immune status were measured in the micro-device. Relative concentrations of lymphocytes and subpopulations of lymphocytes are compared to those obtained with a conventional flow cytometer. The stability of the hydrodynamic focusing and the optical setup was determined by measuring the variation of the signal pulse height of fluorescence calibration beads, being about 2% for the micro-device. This value and the overall performance of the microdevice are similar to conventional flow cytometers. It follows from our results that such microfluidic structures are well suited as modules in a compact, portable read-out instrument. The production process of the microflow cytometers, which we exploited for immunological cell differentiation, is compatible with mass production technology like injection molding and, hence, low cost disposable chips could be available in the future. '

Significance: Fluorescence imaging of rheumatoid diseases with indocyanine green (ICG) is an emerging technique with unique potential for diagnosis and therapy. Device characterization, monitoring of the performance, and further developments of the technique require tissueequivalent fluorescent phantoms of high stability with appropriate anatomical shapes.Aim: Our investigations aim at the development of a three-dimensional (3D) printing technique to fabricate hand and finger models with appropriate optical properties in the near-infrared spectral range. These phantoms should have fluorescence properties similar to ICG, and excellent photostability and durability over years.Approach: We modified a 3D printing methacrylate photopolymer by adding the fluorescent dye Lumogen IR 765 to the raw material. Reduced scattering and absorption coefficients were adjusted to values representative of the human hand by incorporating titanium dioxide powder and black ink. The properties of printed phantoms of various compositions were characterized using UV/Vis and fluorescence spectroscopy, and time-resolved measurements. Photostability and bleaching were investigated with a hand imager. For comparison, several phantoms with ICG as fluorescent dye were printed and characterized as well.Results: The spectral properties of Lumogen IR 765 are very similar to those of ICG. By optimizing the concentrations of Lumogen, titanium dioxide, and ink, anatomically shaped hand and vessel models with properties equivalent to in vivo investigations with a fluorescence hand imager could be printed. Phantoms with Lumogen IR 765 had an excellent photostability over up to 4 years. In contrast, phantoms printed with ICG showed significant bleaching and degradation of fluorescence over time.Conclusions: 3D printing of phantoms with Lumogen IR 765 is a promising method for fabricating anatomically shaped fluorescent tissue models of excellent stability with spectral properties similar to ICG. The phantoms are well-suited to monitor the performance of hand imagers. Concepts can easily be transferred to other fluorescence imaging applications of ICG.

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