Intervertebral Disc-on-a-ChipMF: A New Model for Mouse Disc Culture via Integrating Mechanical Loading and Dynamic Media Flow.

This study aims to develop an ex vivo organ-on-a-chip model, intervertebral Disc-on-a-ChipMF, to investigate integrated effects of mechanical loading and nutrition on disc health. The system consists of a detachable multilayer microfluidic chip, a Computer-Arduino-based control system, and a mechanical loading unit, which were optimized for accurate axial force measurement and the maintenance of a 21-day ex vivo disc culture. To ensure accuracy of axial force, we optimized the axial mechanical loading regimen, used the Computer-Arduino-based system and low-profile force sensors (LPFS) to control the mechanical loading unit, and modeled the force distribution by using computational simulation. A 21-day ex vivo disc culture was demonstrated using the Disc-on-a-ChipMF system, with optimized mechanical loading (0.02 MPa at 1Hz, 1.5 hr/day) and flow rate (1 µL/min). The structural integrity, collagen breakdown, catabolic enzyme activities, and disc cell and collagen alignment revealed that the on-chip cultured discs exhibited a preferred disc health similar to that of native discs for up to 21 days, while discs in a static culture showed detrimental degenerative changes. The mouse Disc-on-a-ChipMF system mimics in vivo disc microenvironment and provides a valuable platform for studying the effects of various factors on disc health and degeneration and testing new therapies.

A three-dimensional microfluidic approach to scaling up microencapsulation of cells.

Current applications of the microencapsulation technique include the use of encapsulated islet cells to treat Type 1 diabetes, and encapsulated hepatocytes for providing temporary but adequate metabolic support to allow spontaneous liver regeneration, or as a bridge to liver transplantation for patients with chronic liver disease. Also, microcapsules can be used for controlled delivery of therapeutic drugs. The two most widely used devices for microencapsulation are the air-syringe pump droplet generator and the electrostatic bead generator, each of which is fitted with a single needle through which droplets of cells suspended in alginate solution are produced and cross-linked into microbeads. A major drawback in the design of these instruments is that they are incapable of producing sufficient numbers of microcapsules in a short-time period to permit mass production of encapsulated and viable cells for transplantation in large animals and humans. We present in this paper a microfluidic approach to scaling up cell and protein encapsulations. The microfluidic chip consists of a 3D air supply and multi-nozzle outlet for microcapsule generation. It has one alginate inlet and one compressed air intlet. The outlet has 8 nozzles, each having 380 micrometers inner diameter, which produce hydrogel microspheres ranging from 500 to 700 µm in diameter. These nozzles are concentrically surrounded by air nozzles with 2 mm inner diameter. There are two tubes connected at the top to allow the air to escape as the alginate solution fills up the chamber. A variable flow pump 115 V is used to pump alginate solution and Tygon® tubing is used to connect in-house air supply to the air channel and peristaltic/syringe pump to the alginate chamber. A pressure regulator is used to control the flow rate of air. We have encapsulated islets and proteins with this high throughput device, which is expected to improve product quality control in microencapsulation of cells, and hence the outcome of their transplantation.

Smartphone-microfluidic fluorescence imaging system for studying islet physiology.

Smartphone technology has been recently applied for biomedical image acquisition and data analysis due to its high-quality imaging capability, and flexibility to customize multi-purpose apps. In this work, we developed and characterized a smartphone-microfluidic fluorescence imaging system for studying the physiology of pancreatic islets. We further evaluated the system capability by performing real-time fluorescence imaging on mouse islets labeled with either chemical fluorescence dyes or genetically encoded fluorescent protein indicators (GEFPIs). Our results showed that the system was capable of analyzing key beta-cell insulin stimulator-release coupling factors in response to various stimuli with high-resolution dynamics. Furthermore, the integration of a microfluidics allowed high-resolution detection of insulin secretion at single islet level. When compared to conventional fluorescence microscopes and macro islet perifusion apparatus, the system has the advantages of low cost, portable, and easy to operate. With all of these features, we envision that this smartphone-microfluidic fluorescence imaging system can be applied to study islet physiology and clinical applications.

A Smartphone-Fluidic Digital Imaging Analysis System for Pancreatic Islet Mass Quantification.

Islet beta-cell viability, function, and mass are three decisive attributes that determine the efficacy of human islet transplantation for type 1 diabetes mellitus (T1DM) patients. Islet mass is commonly assessed manually, which often leads to error and bias. Digital imaging analysis (DIA) system has shown its potential as an alternative, but it has some associated limitations. In this study, a Smartphone-Fluidic Digital Imaging Analysis (SFDIA) System, which incorporates microfluidic techniques and Python-based video processing software, was developed for islet mass assessment. We quantified islets by tracking multiple moving islets in a microfluidic channel using the SFDIA system, and we achieved a relatively consistent result. The counts from the SFDIA and manual counting showed an average difference of 2.91 ± 1.50%. Furthermore, our software can analyze and extract key human islet mass parameters, including quantity, size, volume, IEq, morphology, and purity, which are not fully obtainable from traditional manual counting methods. Using SFDIA on a representative islet sample, we measured an average diameter of 99.88 ± 53.91 µm, an average circularity of 0.591 ± 0.133, and an average solidity of 0.853 ± 0.107. Via analysis of dithizone-stained islets using SFDIA, we found that a higher islet tissue percentage is associated with top-layer islets as opposed to middle-layer islets (0.735 ± 0.213 and 0.576 ± 0.223, respectively). Our results indicate that the SFDIA system can potentially be used as a multi-parameter islet mass assay that is superior in accuracy and consistency, when compared to conventional manual techniques.

A multi-throughput mechanical loading system for mouse intervertebral disc.

Mechanical loading plays an important role in maintaining disc health and function, and in particular, excessive mechanical loading has been identified as one of major reasons of disc degeneration. Intervertebral disc organ culture serves as a valuable tool to study disc biology/pathology. In this study, we report the development and validation of a new mouse disc organ culture system by dynamically applying compression loading in a customized micro-culture device tailored for mouse lumbar discs. Precise axial compression force was delivered by a computer-controlled system consisting of a robust micromechanical linear actuator, a force sensitive resistor, and a precision micro-stepping machinery. Customized PDMS-based loading chambers allowed simultaneous loading of six discs per regimen, which streamlined the workflow to reach sufficient statistic power. The detrimental loading regimen of mouse lumbar discs (0.5 MPa of axial compression at 1Hz for 7 days) was demonstrated through live-dead assay, histology, and fluorescence probe based collagen staining. In addition, various mechanical compression profiles were simulated using different materials and geometry designs, potentiating for more sophisticated loading protocols. In summary, we developed a new mechanical loading system for dynamic axial compression of mouse discs, which created a unique avenue to study disc pathogenesis with enriched mouse species-related resources, and complemented the existing spectrum of bioreactor systems predominately for discs of human and large animals.

An integrated fiberoptic-microfluidic device for agglutination detection and blood typing.

In this paper, an integrated fiberoptic-microfluidic device for the detection of agglutination for blood type cross-matching has been described. The device consists of a straight microfluidic channel through with a reacted RBC suspension is pumped with the help of a syringe pump. The flow intersects an optical path created by an emitter-received fiber optic pair integrated into the microfluidic device. A 650 nm laser diode is used as the light source and a silicon photodiode is used to detect the light intensity. The spacing between the tips of the two optic fibers can be adjusted. When fiber spacing is large and the concentration of the suspension is high, scattering phenomenon becomes the dominant mechanism for agglutination detection while at low concentrations and small spacing, optointerruption becomes the dominant mechanism. An agglutination strength factor (ASF) is calculated from the data. Studies with a variety of blood types indicate that the sensing method correctly identifies the agglutination reaction in all cases. A disposable integrated device can be designed for future implementation of the method for near-bedside pre-transfusion check.

A capacitive displacement sensing technique for early detection of unbalanced loads in a washing machine.

Horizontal axis washing machines are water and energy efficient and becoming popular in the USA. Unlike a vertical axis washer, these do not have an agitator and depend solely on tumbling for the agitation of laundry during the wash cycle. However, due to the constant shifting of laundry during washing, the load distribution is often unbalanced during the high speed spin cycle. We present a displacement-based sensing method to detect unbalance early while the spin rate (rpm) is well below the resonance frequency so that corrective actions may be taken prior to the high speed spin cycle. Experimental and analytical characterizations of the sensor configuration are presented. Results show that the displacement sensor is more appropriate than an accelerometer for this application and offer the potential for a simple, reliable, low cost detection of unbalance.

Microfluidic Approach to Cell Microencapsulation.

Bioartificial pancreas made of insulin-secreting islets cells holds great promise in the treatment of individuals with Type-1 diabetes. Successful islet cell microencapsulation in biopolymers is a key step for providing immunoisolation of transplanted islet cells. Because of the variability in the size and shape of pancreatic islets, one of the main obstacles in their microencapsulation is the inability to consistently control shape, size, and microstructure of the encapsulating biopolymer capsule. In this chapter, we provide a detailed description of a microfluidic approach to islet cell encapsulation in alginate that might address the microencapsulation challenges.