3D-Printed Bubble-Free Perfusion Cartridge System for Live-Cell Imaging

Abstract

The advent of 3D-printing technologies has had a significant effect on the development of medical and biological devices. Perfusion chambers are widely used for live-cell imaging in cell biology research; however, air-bubble invasion is a pervasive problem in perfusion systems. Although 3D printing allows the rapid fabrication of millifluidic and microfluidic devices with high resolution, little has been reported on 3D-printed fluidic devices with bubble trapping systems. Herein, we present a 3D-printed millifluidic cartridge system with bent and flat tapered flow channels for preventing air-bubble invasion, irrespective of bubble volume and without the need for additional bubble-removing devices. This system realizes bubble-free perfusion with a user-friendly interface and no-time-penalty manufacturing processes. We demonstrated the bubble removal capability of the cartridge by continually introducing air bubbles with different volumes during the calcium imaging of Sf21 cells expressing insect odorant receptors. Calcium imaging was conducted using a low-magnification objective lens to show the versatility of the cartridge for wide-area observation. We verified that the cartridge could be used as a chemical reaction chamber by conducting protein staining experiments. Our cartridge system is advantageous for a wide range of cell-based bioassays and bioanalytical studies, and can be easily integrated into portable biosensors.

3D Printing Technologies in Biosensors Production: Recent Developments

“…Material jetting can rely on its ability to print different objects in terms of mechanical properties with high resolution, even though it has its own limitation regarding printing time and accessibility [80,81]. In recent years, material jetting technology has been adopted to integrate 3D-printed microfluidics with biosensors, achieving high resolution (range of tens of microns), resulting in surface roughness of the printed device, its deformation, and resistance to harsh conditions [52][53][54][55]. Microfluidics, in fact, refers to the applied science of high-precision handling of fluids that are geometrically forced into networks of small channels, usually with micro-meter scale dimensions.…”

“…These results lay the foundation for the future use of 3D-printed microfluidic devices for the detection of tumor cell DNA, coupling this highly efficient capture ability with DNA-based detection techniques, such as polymerase chain reaction (PCR) and identification (DNA sequencing) in clinical diagnosis and cancer treatment [52]. Terutsuki et al [53] employed a MultiJet printer in its maximum resolution mode for cartridge production, employing a curable and biocompatible acrylic resin as printing material. The printing process was relatively fast (approximately 4 h).…”

“…A bubble-free perfusion with a user-friendly interface and no-time-penalty manufacturing process was implemented, and the cartridge system proved to be advantageous for different cell-based bioassays and bioanalytical studies. Additionally, it can be integrated into portable biosensors, expanding their spectrum of use [53]. MJP technology was also applied by Graham et al [54] to develop an integrated system that coupled microfluidics and silicon porous nanostructure-based aptasensors for label-free protein detection.…”

Permeation-Enhanced Degassing Method Based on Xylem Embolism Repair and Gas Permeable Materials

“…These bubbles can block the microchannels, decrease the sensitivity of devices, cause inaccuracy of analysis results, and even damage the functional region. For instance, bubbles can lead to nonuniform thermal distribution in the polymerase chain reaction (PCR) chamber, 1,2 reduce cell viability in cell culture devices, 3,4 and cause nonuniform cell loading in drug screening microfluidic devices. 5 In previous reports, several pretreating degassing methods have been proposed to eliminate bubbles in microchannels, including the method of sealing the device's outlet temporarily while increasing the inlet pressure simultaneously, 6 the method of flushing microchannels with low-polarity aqueous solutions 7,8 and the method of integrating microstructures in the device aiming at reducing the possibility of bubble formation.…”

“…These bubbles can block the microchannels, decrease the sensitivity of devices, cause inaccuracy of analysis results, and even damage the functional region. For instance, bubbles can lead to nonuniform thermal distribution in the polymerase chain reaction (PCR) chamber, , reduce cell viability in cell culture devices, , and cause nonuniform cell loading in drug screening microfluidic devices …”