Centrifugal Microfluidic Systems Research Articles

Flipping: A Valve-Free Strategy to Control Fluid Flow in Centrifugal Microfluidic Systems

Centrifugal microfluidics has emerged as a technology to automate fluid handling at the microliter scale in bioassays. It has been mostly developed for point-of-care diagnostic applications, where assays involve a well-defined workflow. By contrast, this study introduces a microfluidic strategy that allows multiple workflows within the same device, thereby addressing the requirements of sample preparation steps in R&D settings. The corresponding device is a microfluidic chip that can be inserted either in a lab-on-a-disk or in the swinging-bucket of a commercial centrifuge. The control of liquid transfers from one chamber to the next is achieved by flipping the chip, thereby reorienting it in the centrifugal force field. The present experimental study describes the fluid instability that triggers the transfer upon flipping. A semi-empirical physical model is developed that predicts the acceleration threshold of the instability as a function of design parameters. By contrast to most existing control strategies in centrifugal microfluidics, the present strategy does not involve any valve. Consequently, it is robust to variations of material and sample properties, and it yields minimal fabrication constraints. Designing appropriately sized chambers enables flexible implementation of complex workflows. This strategy brings the miniaturization benefits of centrifugal microfluidics to sample preparation workflows.

Digital process control of multi-step assays on centrifugal platforms using high-low-high rotational-pulse triggered valving.

Due to their capability for comprehensive sample-to-answer automation, the interest in centrifugal microfluidic systems has greatly increased in industry and academia over the last quarter century. The main applications of these "Lab-on-a-Disc" (LoaD) platforms are in decentralised bioanalytical point-of-use / point-of-care testing. Due to the unidirectional and omnipresent nature of the centrifugal force, advanced flow control is key to coordinate multi-step / multi-reagent assay formats on the LoaD. Formerly, flow control was often achieved by capillary burst valves which require gradual increments of the spin speed of the system-innate spindle motor. Recent advanced introduced a flow control scheme called 'rotational pulse actuated valves'. In these valves the sequence of valve actuation is determined by the architecture of the disc while actuation is triggered by freely programmable upward spike (i.e. Low-High-Low (LHL)) in the rotational frequency. This paradigm shift from conventional 'analogue' burst valves to 'digital' pulsing significantly increases the number of sequential while also improving the overall robustness of flow control. In this work, we expand on these LHL valves by introducing High-Low-High (HLH) pulse-actuated (PA) valving which are actuated by 'downward' spike in the disc spin-rate. These HLH valves are particularly useful for high spin-rate operations such as centrifugation of blood. We introduce two different HLH architectures and then combine the most promising with LHL valves to implement the time-dependent liquid handling protocol underlying a common liver function test panel.

Recent Developments in Inertial and Centrifugal Microfluidic Systems along with the Involved Forces for Cancer Cell Separation: A Review.

The treatment of cancers is a significant challenge in the healthcare context today. Spreading circulating tumor cells (CTCs) throughout the body will eventually lead to cancer metastasis and produce new tumors near the healthy tissues. Therefore, separating these invading cells and extracting cues from them is extremely important for determining the rate of cancer progression inside the body and for the development of individualized treatments, especially at the beginning of the metastasis process. The continuous and fast separation of CTCs has recently been achieved using numerous separation techniques, some of which involve multiple high-level operational protocols. Although a simple blood test can detect the presence of CTCs in the blood circulation system, the detection is still restricted due to the scarcity and heterogeneity of CTCs. The development of more reliable and effective techniques is thus highly desired. The technology of microfluidic devices is promising among many other bio-chemical and bio-physical technologies. This paper reviews recent developments in the two types of microfluidic devices, which are based on the size and/or density of cells, for separating cancer cells. The goal of this review is to identify knowledge or technology gaps and to suggest future works.

Modular design of centrifugal microfluidic system and its application in nucleic acid screening

Modular integration of functional components on the chip and increasement in control accuracy through real-time alteration in the force direction of droplets is an effective way to optimize centrifugal microfluidic systems and realize passive components, compact modules, and high-throughput control. Conventional centrifugal microfluidic chips are mainly driven and controlled by centrifugal force and Euler force. The control valves are easily affected by machining precision, making the control unstable. In this study, a novel centrifugal microfluidic system is introduced to improve the freedom and accuracy of chip control while facilitating the design and addition of passive functional components. Furthermore, we modularize the centrifugal microfluidic chip to greatly shorten the period of design and optimization cycle and achieve chip reusability and multi-threaded control. Finally, to verify the feasibility of the modular centrifugal microfluidic chip applied to high-throughput nucleic acid screening, we test the nucleic acid purification and detection colorimetric reactions based on the modular centrifugal microfluidic chip. Among them, Chelex-100 is used to realize the purification of nucleic acid in cell lysate, and the purified solution can realize amplification in the PCR instrument, and the nucleic acid detection results are consistent with the off-chip kit by experimental testing. The system has great flexibility and stability under the acceptable purity of nucleic acid, which indicates that the platform has great potential for large-scale rapid screening applications.

Plant pathogen detection on a lab-on-a-disc using solid-phase extraction and isothermal nucleic acid amplification enabled by digital pulse-actuated dissolvable film valves

By virtue of its ruggedness, portability, rapid processing times, and ease-of-use, academic and commercial interest in centrifugal microfluidic systems has soared over the last decade. A key advantage of the LoaD platform is the ability to automate laboratory unit operations (LUOs) (mixing, metering, washing etc.) to support direct translation of ‘on-bench’ assays to ‘on-chip’. Additionally, the LoaD requires just a low-cost spindle motor rather than specialized and expensive microfluidic pumps. Furthermore, when flow control (valves) is implemented through purely rotational changes in this same spindle motor (rather than using additional support instrumentation), the LoaD offers the potential to be a truly portable, low-cost and accessible platform. Current rotationally controlled valves are typically opened by sequentially increasing the disc spin-rate to a specific opening frequency. However, due lack of manufacturing fidelity these specific opening frequencies are better described as spin frequency ‘bands’. With low-cost motors typically having a maximum spin-rate of 6000 rpm (100 Hz), using this ‘analogue’ approach places a limitation on the number of valves, which can be serially actuated thus limiting the number of LUOs that can be automated. In this work, a novel flow control scheme is presented where the sequence of valve actuation is determined by architecture of the disc while its timing is governed by freely programmable ‘digital’ pulses in its spin profile. This paradigm shift to ‘digital’ flow control enables automation of multi-step assays with high reliability, with full temporal control, and with the number of LUOs theoretically only limited by available space on the disc. We first describe the operational principle of these valves followed by a demonstration of the capability of these valves to automate complex assays by screening tomato leaf samples against plant pathogens. Reagents and lysed sample are loaded on-disc and then, in a fully autonomous fashion using only spindle-motor control, the complete assay is automated. Amplification and fluorescent acquisition take place on a custom spin-stand enabling the generation of real-time LAMP amplification curves using custom software. To prevent environmental contamination, the entire discs are sealed from atmosphere following loading with internal venting channels permitting easy movement of liquids about the disc. The disc was successfully used to detect the presence of thermally inactivated Clavibacter michiganensis. Michiganensis (CMM) bacterial pathogen on tomato leaf samples.

Active pumping and control of flows in centrifugal microfluidics

This review is an account of centrifugal microfluidic systems that use various actuation strategies in addition to intrinsic centrifugal forces to accurately regulate the motion of fluids during rotation. Platforms that integrate active methods of pumping and flow control render centrifugal microfluidics more versatile as they facilitate integration and process automation by enabling (or improving the reliability of) important fluidic functions, such as metering, aliquoting, valving, flow switching, mixing, and inward pumping. Principles and working mechanisms underlying these strategies are described in the context of recent trends in instrument design and development where centrifugal platforms have been equipped with pneumatic, magnetic or electromechanical actuators serving as pumping and valving elements. The potential of these platforms to perform complex bioanalytical assays in an automated fashion is illustrated by several examples, which include on-chip preparation of aliquot libraries, nucleic acid purification, amplification and analysis as well as blood separation.

Design and fabrication of a centrifugal microfluidic disc including septum valve for measuring hemoglobin A1c in human whole blood using immunoturbidimetry method

Diabetes mellitus is a global endemic with a rapidly increasing prevalence in both developing and developed countries. Recently, hemoglobin A1c has been recommended by the American Diabetes Associations as a possible substitute for fasting blood glucose for the diagnosis of diabetes, because it is an indicator of long-term glycemic control. Also, centrifugal microfluidic systems have good potential for use in the point of care testing systems. In this study, a centrifugal microfluidic disc was designed and manufactured to measure hemoglobin A1c in whole blood using an immunoturbidimetry based method. Also, a new passive valve, named septum valve, was presented to precisely control the entry and exit of reagents. This design comprises three inputs for injection of reagents and a blood sample, three septum valves, a two-part mixing chamber and a chamber to measure the absorbance of the sample based on the immunoturbidimetry method. Fourteen blood samples were tested using the manufactured disc, and the results were very congruous with the clinical data. By using the designed centrifugal microfluidic disc, the hemoglobin A1c in whole blood was measured in 8 min, with a standard deviation of ± 0.36% HbA1c.

Wedge actuated normally-open and normally-closed valves for centrifugal microfluidic applications

Normally-open and normally-closed valves are difficult to achieve and highly demanded in centrifugal microfluidic systems. This paper reports a novel pinch-type valving technology that can be used to realize both normally-closed and normally-open valves for fluidic control on centrifugal microfluidic platform. A sliding wedge with inclined surface planes was used to transduce the actuation of the centrifugal force from radial direction to the perpendicular direction of the microfluidic disc. The function of the valves has been demonstrated experimentally using a PDMS microfluidic system. The burst frequency of normally-closed valves can be tuned through changing the physical parameters of the valving system. The normally-open valve shows good performance at blocking the flow at high rotation speed and maintained blocking of the sample flow until the platform completely stops. The valving technology offers key advantages over current active valving techniques for Lab-on-CD, including simpler actuation process, less contamination to samples, better reversibility, and more effective in implementing of normally-open valves.

Baking Powder Actuated Centrifugo-Pneumatic Valving for Automation of Multi-Step Bioassays.

We report a new flow control method for centrifugal microfluidic systems; CO2 is released from on-board stored baking powder upon contact with an ancillary liquid. The elevated pressure generated drives the sample into a dead-end pneumatic chamber sealed by a dissolvable film (DF). This liquid incursion wets and dissolves the DF, thus opening the valve. The activation pressure of the DF valve can be tuned by the geometry of the channel upstream of the DF membrane. Through pneumatic coupling with properly dimensioned disc architecture, we established serial cascading of valves, even at a constant spin rate. Similarly, we demonstrate sequential actuation of valves by dividing the disc into a number of distinct pneumatic chambers (separated by DF membranes). Opening these DFs, typically through arrival of a liquid to that location on a disc, permits pressurization of these chambers. This barrier-based scheme provides robust and strictly ordered valve actuation, which is demonstrated by the automation of a multi-step/multi-reagent DNA-based hybridization assay.

A numerical study of the Coriolis effect in centrifugal microfluidics with different channel arrangements

The Coriolis force has been of great interest to control the transversal flow that is critical for mixing or switching fluids in centrifugal microfluidics. Therefore, the variation in the Coriolis effect has been extensively investigated by varying the rotational speeds and the cross-sectional geometry of microchannels. However, the subject of such investigations has been limited to radially positioned microchannels even though channels can lie everywhere on the rotating platform with different arrangements. In this study, we use numerical methods to investigate the Coriolis effect resulting from the angular alignment (AA) and radial displacement (RD) of rotating microchannels. Our analysis focuses on determining the contribution that different channel arrangements have in the deviation of parabolic velocity profiles, which is generally produced by the effective Coriolis force. We found that the flow can deviate even at a low rotational speed, where the Coriolis force is negligible, with an AA of up to 33 % which is a sufficient amount for flow switching. Once the rotational speed reaches to the critical RPM, the flow deviates by an effective Coriolis force, but the deviation systematically varies with AA or RD. As the Coriolis force becomes more dominant with a high rotational speed, the deviation reaches a saturation point, while flow rate is regulated by AA or RD, enabling the flow rate to remain low even at very high RPM, without reducing the deviation. The variation in the Coriolis effect due to the different channel arrangements investigated in this study is believed to provide an essential basis to design and develop centrifugal microfluidic systems.

Challenges and Opportunities of Centrifugal Microfluidics for Extreme Point-of-Care Testing.

The advantages offered by centrifugal microfluidic systems have encouraged its rapid adaptation in the fields of in vitro diagnostics, clinical chemistry, immunoassays, and nucleic acid tests. Centrifugal microfluidic devices are currently used in both clinical and point-of-care settings. Recent studies have shown that this new diagnostic platform could be potentially used in extreme point-of-care settings like remote villages in the Indian subcontinent and in Africa. Several technological inventions have decentralized diagnostics in developing countries; however, very few microfluidic technologies have been successful in meeting the demand. By identifying the finest difference between the point-of-care testing and extreme point-of-care infrastructure, this review captures the evolving diagnostic needs of developing countries paired with infrastructural challenges with technological hurdles to healthcare delivery in extreme point-of-care settings. In particular, the requirements for making centrifugal diagnostic devices viable in developing countries are discussed based on a detailed analysis of the demands in different clinical settings including the distinctive needs of extreme point-of-care settings.

CD-Based Microfluidics for Primary Care in Extreme Point-of-Care Settings.

We review the utility of centrifugal microfluidic technologies applied to point-of-care diagnosis in extremely under-resourced environments. The various challenges faced in these settings are showcased, using areas in India and Africa as examples. Measures for the ability of integrated devices to effectively address point-of-care challenges are highlighted, and centrifugal, often termed CD-based microfluidic technologies, technologies are presented as a promising platform to address these challenges. We describe the advantages of centrifugal liquid handling, as well as the ability of a standard CD player to perform a number of common laboratory tests, fulfilling the role of an integrated lab-on-a-CD. Innovative centrifugal approaches for point-of-care in extremely resource-poor settings are highlighted, including sensing and detection strategies, smart power sources and biomimetic inspiration for environmental control. The evolution of centrifugal microfluidics, along with examples of commercial and advanced prototype centrifugal microfluidic systems, is presented, illustrating the success of deployment at the point-of-care. A close fit of emerging centrifugal systems to address a critical panel of tests for under-resourced clinic settings, formulated by medical experts, is demonstrated. This emphasizes the potential of centrifugal microfluidic technologies to be applied effectively to extremely challenging point-of-care scenarios and in playing a role in improving primary care in resource-limited settings across the developing world.

The use of polybutene for controlling the flow of liquids in centrifugal microfluidic systems

The field of centrifugal microfluidics has evolved over the last several decades to allow implementation of complex biological and chemical assays on Lab-on-Disc (LOD) platforms. Present study describes the use of polymer polybutene for tuning hydrophobic siphons and for liquid volume definition on a centrifugal microfluidic platform. Both the siphon tuning and the volume definition steps are carried out by generating negative pressure that results from the volume expansion caused by the transfer of polybutene from a dedicated chamber into a secondary reservoir via a connecting siphon. The hydrophobic valve of the chamber that holds polybutene bursts open at specific angular velocities that depend on the height and density of the liquid column. Thus, the parameters of siphon activation can be adjusted by tuning the burst angular velocity of the valve that is driven by filling the tuning reservoir with a specific volume of polybutene. The same disc construction can be utilized to provide volume definition functionality to transfer liquids from one reservoir to another reservoir in as many fractions as there are immiscible liquids of different densities in the tuning chamber. The presented work also demonstrates the use of polybutene in sealing fluidic chambers to improve heating efficiency and to minimize evaporation during thermal cycling required for applications such as PCR amplification. Finally, the use of polybutene as a stationary liquid phase in droplet production on a spinning disc is demonstrated.

Reversible thermo-pneumatic valves on centrifugal microfluidic platforms.

Centrifugal microfluidic systems utilize a conventional spindle motor to automate parallel biochemical assays on a single microfluidic disk. The integration of complex, sequential microfluidic procedures on these platforms relies on robust valving techniques that allow for the precise control and manipulation of fluid flow. The ability of valves to consistently return to their former conditions after each actuation plays a significant role in the real-time manipulation of fluidic operations. In this paper, we introduce an active valving technique that operates based on the deflection of a latex film with the potential for real-time flow manipulation in a wide range of operational spinning speeds. The reversible thermo-pneumatic valve (RTPV) seals or reopens an inlet when a trapped air volume is heated or cooled, respectively. The RTPV is a gas-impermeable valve composed of an air chamber enclosed by a latex membrane and a specially designed liquid transition chamber that enables the efficient usage of the applied thermal energy. Inputting thermo-pneumatic (TP) energy into the air chamber deflects the membrane into the liquid transition chamber against an inlet, sealing it and thus preventing fluid flow. From this point, a centrifugal pressure higher than the induced TP pressure in the air chamber reopens the fluid pathway. The behaviour of this newly introduced reversible valving system on a microfluidic disk is studied experimentally and theoretically over a range of rotational frequencies from 700 RPM to 2500 RPM. Furthermore, adding a physical component (e.g., a hemispherical rubber element) to induce initial flow resistance shifts the operational range of rotational frequencies of the RTPV to more than 6000 RPM. An analytical solution for the cooling of a heated RTPV on a spinning disk is also presented, which highlights the need for the future development of time-programmable RTPVs. Moreover, the reversibility and gas impermeability of the RTPV in the microfluidic networks are validated on a microfluidic disk designed for performing liquid circulation. Finally, an array of RTPVs is integrated into a microfluidic cartridge to enable sequential aliquoting for the conversion of dengue virus RNA to cDNA and the preparation of PCR reaction mixtures.

Rapid prototyping of pneumatically actuated hydrocarbon gel valves for centrifugal microfluidic devices

A novel, easy to prototype hydrocarbon gel-based active valve was developed for use in centrifugal microfluidic devices. The valve has been demonstrated to restrict flow by an additional 1000 revolutions per minute (RPM) when compared to a passive capillary valve of the same size located at the same radius. Opening of the valve is accomplished in a contactless manner using a stream of focused compressed air. The ease of fabrication, low cost and small dimensions of the gel valve offer the potential for integration of multiple valves of this type into multi-process centrifugal microfluidic systems.

The Effect of Contact Angles and Capillary Dimensions on the Burst Frequency of Super Hydrophilic and Hydrophilic Centrifugal Microfluidic Platforms, a CFD Study

This paper employs the volume of fluid (VOF) method to numerically investigate the effect of the width, height, and contact angles on burst frequencies of super hydrophilic and hydrophilic capillary valves in centrifugal microfluidic systems. Existing experimental results in the literature have been used to validate the implementation of the numerical method. The performance of capillary valves in the rectangular and the circular microfluidic structures on super hydrophilic centrifugal microfluidic platforms is studied. The numerical results are also compared with the existing theoretical models and the differences are discussed. Our experimental and computed results show a minimum burst frequency occurring at square capillaries and this result is useful for designing and developing more sophisticated networks of capillary valves. It also predicts that in super hydrophilic microfluidics, the fluid leaks consistently from the capillary valve at low pressures which can disrupt the biomedical procedures in centrifugal microfluidic platforms.

Modeling of Capillary Burst Valve and Siphon with Hydrophilic Cover for Centrifugal Microfluidic Systems

This paper reports numerical and experimental investigation of capillary burst valve and siphon, which were used in the centrifugal microfluidic systems widely. The simulation model is based on the solution of the minimum energy problem by the Surface Evolver software. In contrast to the solution of meniscus equilibrium equations, it can provide the appropriate boundary conditions for the microvalves with a hydrophilic flat cover. As an experimental verification, a centrifugal microfluidic device based on PDMS and glass was fabricated by soft lithography. The simulated results agree well with experimental data. The technology is useful for the design of microfluidic systems with complex wetting characters and geometries.

Centrifugal microfluidics for cell analysis

Over the past two decades, centrifugal microfluidic systems have successfully demonstrated their capability for robust, high-performance liquid handling to enable modular, multi-purpose lab-on-a-chip platforms for a wide range of life-science applications. Beyond the handling of homogeneous liquids, the unique, rotationally controlled centrifugal actuation has proven to be specifically advantageous for performing cell and particle handling and assays. In this review we discuss technologies to implement two important steps for cell handling, namely separation and capturing/counting.

Stacked centrifugal microfluidic device with three-dimensional microchannel networks and multifunctional capillary bundle structures for immunoassay

A stacked-type centrifugal microfluidic device with three-dimensional (3D) microchannel networks and multifunctional capillary bundle structures is developed, and an immunoassay for the detection of immunoglobulin G (IgG) is demonstrated. The device consists of multiple layers of disk-like chips with conventional planar microchannels and vertical capillary bundle structures that are constructed on disks of poly(dimethylsiloxane) and poly(methyl methacrylate). The integration of multiple reactors up to 10 reactors on a disk with 8cm in diameter is realized by reducing the area of reservoir but increasing the thickness. Moreover the thick optical detection reservoir is also realized for absorption spectroscopy in the 3D centrifugal microfluidics. 3D liquid valving on the vertical capillary bundle structures and liquid transportation through these structures are successfully demonstrated. The results show the successful control of the multistep liquid injection without problems such as blockage of capillary bundle by bubbles that expected to be caused due to 3D structure. The results of the immunoassay show clear response to analyte concentration suggesting successful unit operations in developed 3D centrifugal microfluidic device for immunoassay. Finally, the advantages and impact of the 3D design for the development of high-performance centrifugal microfluidic systems are summarized.

Centrifugo-pneumatic valving utilizing dissolvable films

In this article we introduce a novel technology that utilizes specialized water dissolvable thin films for valving in centrifugal microfluidic systems. In previous work (William Meathrel and Cathy Moritz, IVD Technologies, 2007), dissolvable films (DFs) have been assembled in laminar flow devices to form efficient sacrificial valves where DFs simply open by direct contact with liquid. Here, we build on the original DF valving scheme to leverage sophisticated, merely rotationally actuated vapour barriers and flow control for enabling comprehensive assay integration with low-complexity instrumentation on "lab-on-a-disc" platforms. The advanced sacrificial valving function is achieved by creating an inverted gas-liquid stack upstream of the DF during priming of the system. At low rotational speeds, a pocket of trapped air prevents a surface-tension stabilized liquid plug from wetting the DF membrane. However, high-speed rotation disrupts the metastable gas/liquid interface to wet the DF and thus opens the valve. By judicious choice of the radial position and geometry of the valve, the burst frequency can be tuned over a wide range of rotational speeds nearly 10 times greater than those attained by common capillary burst valves based on hydrophobic constrictions. The broad range of reproducible burst frequencies of the DF valves bears the potential for full integration and automation of comprehensive, multi-step biochemical assay protocols. In this report we demonstrate DF valving, discuss the biocompatibility of using the films, and show a potential sequential valving system including the on-demand release of on-board stored liquid reagents, fast centrifugal sedimentation and vigorous mixing; thus providing a viable basis for use in lab-on-a-disc platforms for point-of-care diagnostics and other life science applications.