Patterning Fluid Shear Stress Landscapes with Multiphoton Inner Laser Lithography (MILL) for Live Cell Adhesion and Translocation

Current 3D microfluidic fabrication methods require hours and specialized equipment to fabricate microstructures in a single channel so as to recapitulate mixed (homogenous and heterogeneous) in vivo fluid flow. Inspired by the ancient art form of inside painting, we developed a technique for 3D fabrication of micro-patterned flow channels and mixed in vivo fluid flow in a matter of minutes. We termed this technique Multiphoton Inner Laser Lithography ( MILL ). We further showed that MILL is compatible with both flat and curved channel shapes. MILL recapitulated in vivo tissue topology and 3D fluid flow of the tissue microenvironment, all of which are vital for understanding of how extracellular fluid flow regulates cell function. Cells in MILL capillary tubes response to a variety of in vivo -like laminar flow patterns (homogenous and heterogeneous). Live cells were observed to organize, translocate and adhere along different fluid shear landscapes (0 - 81 dynes/cm2) in real time. Parallel strips of MILL channels were assembled for platelet function tests (~2000 microthrombi per test). The MILL technique heralds a new paradigm where dynamics of in vivo fluid flow can be readily reproduced in minutes on a standard multiphoton imaging microscope and benefit preclinical screening of drug pharmacokinetics. ### Competing Interest Statement The authors have declared no competing interest.