Synthesis and active manipulation of magnetic liquid beads.

We report the fabrication and characterisation of magnetic liquid beads with a solid magnetic shell and liquid core using microfluidic techniques. The liquid beads consist of a fluorinated oil core and a polymer shell with magnetite particles. The beads are generated in a flow-focusing polydimethylsiloxane (PDMS) device and cured by photo polymerisation. We investigated the response of the liquid beads to an external magnetic field by characterising their motion towards a permanent magnet. Magnetic sorting of liquid beads in a channel was achieved with 90% efficiency. The results show that the liquid beads can be controlled magnetically and have potential applications in digital microfluidics including nucleic acid amplification, drug delivery, cell culture, sensing, and tissue engineering. The present paper also discusses the magnetophoretic behaviour of the liquid bead by varying its mass and magnetite concentration in the shell. We also demonstrated the two-dimensional self-assembly of magnetic liquid beads for potential use in digital polymerase chain reaction and digital loop mediated isothermal amplification.

Calcium alginate elastic capsules for microalgal cultivation.

Calcium alginate elastic capsules with a core-shell structure are versatile spherical solid beads that can be produced in large quantities using various techniques. This type of capsule is a promising platform for cell culture applications, owing to its mechanical elasticity and transparency. This paper reports the production of calcium alginate capsules with high consistency, and for the first time, demonstrates the feasibility of the capsules for microalgal cultivation. Cell growth analysis reveals that the vibrationally-shaken calcium alginate elastic capsule platform yielded a higher maximum cell number (4.86 × 108 cells per mL) during the cultivation period than the control solution platforms. Aquafeed and food supplements for humans are the targeted applications of this novel platform.

Recent developments and future perspectives of microfluidics and smart technologies in wearable devices.

Wearable devices are gaining popularity in the fields of health monitoring, diagnosis, and drug delivery. Recent advances in wearable technology have enabled real-time analysis of biofluids such as sweat, interstitial fluid, tears, saliva, wound fluid, and urine. The integration of microfluidics and emerging smart technologies, such as artificial intelligence (AI), machine learning (ML), and Internet of Things (IoT), into wearable devices offers great potential for accurate and non-invasive monitoring and diagnosis. This paper provides an overview of current trends and developments in microfluidics and smart technologies in wearable devices for analyzing body fluids. The paper discusses common microfluidic technologies in wearable devices and the challenges associated with analyzing each type of biofluid. The paper emphasizes the importance of combining smart technologies with microfluidics in wearable devices, and how they can aid diagnosis and therapy. Finally, the paper covers recent applications, trends, and future developments in the context of intelligent microfluidic wearable devices.

Hydrophobically Modified Gelatin Particles for Production of Liquid Marbles.

The unique properties and morphology of liquid marbles (LMs) make them potentially useful for various applications. Non-edible hydrophobic organic polymer particles are widely used to prepare LMs. It is necessary to increase the variety of LM particles to extend their use into food and pharmaceuticals. Herein, we focus on hydrophobically modified gelatin (HMG) as a base material for the particles. The surface tension of HMG decreased as the length of alkyl chains incorporated into the gelatin and the degree of substitution (DS) of the alkyl chains increased. HMG with a surface tension of less than 37.5 mN/m (determined using equations based on the Young-Dupré equation and Kaelble-Uy theory) successfully formed LMs of water. The minimum surface tension of a liquid in which it was possible to form LMs using HMG particles was approximately 53 mN/m. We also showed that the liquid-over-solid spreading coefficient SL/S is a potential new factor for predicting if particles can form LMs. The HMG particles and the new system for predicting LM formation could expand the use of LMs in food and pharmaceuticals.

Lab-on-a-Chip Technologies for Microgravity Simulation and Space Applications.

Gravity plays an important role in the development of life on earth. The effect of gravity on living organisms can be investigated by controlling the magnitude of gravity. Most reduced gravity experiments are conducted on the Lower Earth Orbit (LEO) in the International Space Station (ISS). However, running experiments in ISS face challenges such as high cost, extreme condition, lack of direct accessibility, and long waiting period. Therefore, researchers have developed various ground-based devices and methods to perform reduced gravity experiments. However, the advantage of space conditions for developing new drugs, vaccines, and chemical applications requires more attention and new research. Advancements in conventional methods and the development of new methods are necessary to fulfil these demands. The advantages of Lab-on-a-Chip (LOC) devices make them an attractive option for simulating microgravity. This paper briefly reviews the advancement of LOC technologies for simulating microgravity in an earth-based laboratory.

A Portable Device for LAMP Based Detection of SARS-CoV-2.

This paper reports the design, development, and testing of a novel, yet simple and low-cost portable device for the rapid detection of SARS-CoV-2. The device performs loop mediated isothermal amplification (LAMP) and provides visually distinguishable images of the fluorescence emitted from the samples. The device utilises an aluminium block embedded with a cartridge heater for isothermal heating of the sample and a single-board computer and camera for fluorescence detection. The device demonstrates promising results within 20 min using clinically relevant starting concentrations of the synthetic template. Time-to-signal data for this device are considerably lower compared to standard quantitative Polymerase Chain Reaction(qPCR) machine (~10-20 min vs. >38 min) for 1 × 102 starting template copy number. The device in its fully optimized and characterized state can potentially be used as simple to operate, rapid, sensitive, and inexpensive platform for population screening as well as point-of-need severe acute respiratory syndrome-coronavirus 2 (SARS-CoV-2) detection and patient management.

Loop-Mediated Isothermal Amplification in a Core-Shell Bead Assay for the Detection of Tyrosine Kinase AXL Overexpression.

The upregulated expression of tyrosine kinase AXL has been reported in several hematologic and solid human tumors, including gastric, breast, colorectal, prostate and ovarian cancers. Thus, AXL can potentially serve as a diagnostic and prognostic biomarker for various cancers. This paper reports the first ever loop-mediated isothermal amplification (LAMP) in a core-shell bead assay for the detection of AXL gene overexpression. We demonstrated simple instrumentation toward a point-of-care device to perform LAMP. This paper also reports the first ever use of core-shell beads as a microreactor to perform LAMP as an attempt to promote environmentally-friendly laboratory practices.

A novel RdRp-based colorimetric RT-LAMP assay for rapid and sensitive detection of SARS-CoV-2 in clinical and sewage samples from Pakistan.

Novel corona virus SARS-CoV-2, causing coronavirus disease 2019 (COVID-19), has become a global health challenge particularly for developing countries like Pakistan where overcrowded cities, inadequate sanitation, little health awareness and poor socioeconomic conditions exist. The SARS-CoV-2 has been known to spread primarily through direct contact and respiratory droplets. However, detection of SARS-CoV-2 in stool and sewage have raised the possibility of fecal-oral mode of transmission. Currently, quantitative reverse-transcriptase PCR (qRT-PCR) is the only method being used for SARS-CoV-2 detection, which requires expensive instrumentation, dedicated laboratory setup, highly skilled staff, and several hours to report results. Considering the high transmissibility and rapid spread, a robust, sensitive, specific and cheaper assay for rapid SARS-CoV-2 detection is highly needed. Herein, we report a novel colorimetric RT-LAMP assay for naked-eye detection of SARS-COV-2 in clinical as well as sewage samples. Our SARS-CoV-2 RdRp-based LAMP assay could successfully detect the virus RNA in 26/28 (93%) of RT-PCR positive COVID-19 clinical samples with 100% specificity (n = 7) within 20 min. We also tested the effect of various additives on the performance of LAMP assay and found that addition of 1 mg/ml bovine serum albumin (BSA) could increase the sensitivity of assay up to 101 copies of target sequence. Moreover, we also successfully applied this assay to detect SARS-CoV-2 in sewage waters collected from those areas of Lahore, a city of Punjab province of Pakistan, declared as virus hotspots by local government. Our optimized LAMP assay could provide a sensitive first tier strategy for SARS-CoV-2 screening and can potentially help diagnostic laboratories in better handling of high sample turnout during pandemic situation. By providing rapid naked-eye SARS-CoV-2 detection in sewage samples, this assay may support pandemic readiness and emergency response to any possible virus outbreaks in future.

Correction: Liquid marble-based digital microfluidics - fundamentals and applications.

Correction for 'Liquid marble-based digital microfluidics - fundamentals and applications' by Chin Hong Ooi et al., Lab Chip, 2021, DOI: .

Liquid marble-based digital microfluidics - fundamentals and applications.

Liquid marbles are droplets with volume typically on the order of microliters coated with hydrophobic powder. Their versatility, ease of use and low cost make liquid marbles an attractive platform for digital microfluidics. This paper provides the state of the art of discoveries in the physics of liquid marbles and their practical applications. The paper first discusses the fundamental properties of liquid marbles, followed by the summary of different techniques for the synthesis of liquid marbles. Next, manipulation techniques for handling liquid marbles are discussed. Applications of liquid marbles are categorised according to their use as chemical and biological reactors. The paper concludes with perspectives on the future development of liquid marble-based digital microfluidics.

Detection of the SARS-CoV-2 humanized antibody with paper-based ELISA.

This work reports the development of a rapid, simple and inexpensive colorimetric paper-based assay for the detection of the severe acute respiratory symptom coronavirus 2 (SARS-CoV-2) humanized antibody. The paper device was prepared with lamination for easy sample handling and coated with the recombinant SARS-CoV-2 nucleocapsid antigen. This assay employed a colorimetric reaction, which is followed by horseradish peroxidase (HRP) conjugated detecting antibody in the presence of the 3,3',5,5'-tetramethylbenzidine (TMB) substrate. The colorimetric readout was evaluated and quantified for specificity and sensitivity. The characterization of this assay includes determining the linear regression curve, the limit of detection (LOD), the repeatability, and testing complex biological samples. We found that the LOD of the assay was 9.00 ng µL-1 (0.112 IU mL-1). The relative standard deviation was approximately 10% for a sample number of n = 3. We believe that our proof-of-concept assay has the potential to be developed for clinical screening of the SARS-CoV-2 humanized antibody as a tool to confirm infected active cases or to confirm SARS-CoV-2 immune cases during the process of vaccine development.

Core-Shell Beads as Microreactors for Phylogrouping of E. coli Strains.

Multiplex polymerase chain reaction (PCR) is an effective tool for simultaneous detection of target genes. Nevertheless, their use has been restricted due to the intrinsic interference between primer pairs. Performing several single PCRs in an array format instead of a multiplex PCR is a simple way to overcome this obstacle. However, there are still major technical challenges in designing a new generation of single PCR microreactors with a small sample volume, rapid thermal cycling, and no evaporation during amplification. We report a simple and robust core-shell bead array for a series of single amplifications. Four core-shell beads with a polymer coating and PCR mixture were synthesized using liquid marble formation and subsequent photo polymerization. Each bead can detect one target gene. We constructed a customised system for thermal cycling of these core-shell beads. Phylogrouping of the E. coli strains was carried out based on the fluorescent signal of the core-shell beads. This platform can be a promising alternative for multiplex nucleic acid analyses due to its simplicity and high throughput. The platform reported here also reduces the cycling time and avoids evaporation as well as contamination of the sample during the amplification process.

Core-Shell Beads Made by Composite Liquid Marble Technology as A Versatile Microreactor for Polymerase Chain Reaction.

Over the last three decades, the protocols and procedures of the DNA amplification technique, polymerase chain reaction (PCR), have been optimized and well developed. However, there have been no significant innovations in processes for sample dispersion for PCR that have reduced the amount of single-use or unrecyclable plastic waste produced. To address the issue of plastic waste, this paper reports the synthesis and successful use of a core-shell bead microreactor using photopolymerization of a composite liquid marble as a dispersion process. This platform uses the core-shell bead as a simple and effective sample dispersion medium that significantly reduces plastic waste generated compared to conventional PCR processes. Other improvements over conventional PCR processes of the novel dispersion platform include increasing the throughput capability, enhancing the performance and portability of the thermal cycler, and allowing for the contamination-free storage of samples after thermal cycling.

Liquid marbles as biochemical reactors for the polymerase chain reaction.

The polymerase chain reaction (PCR) is a popular and well-established DNA amplification technique. Technological and engineering advancements in the field of microfluidics have fuelled the progress of polymerase chain reaction (PCR) technology in the last three decades. Advances in microfluidics-based PCR technology have significantly reduced the sample volume and thermal cycling time. Further advances led to novel and accurate techniques such as the digital PCR. However, contamination of PCR samples, lack of reusability of the microfluidic PCR platforms, complexity in instrumentation and operation remain as some of the significant drawbacks of conventional microfluidic PCR platforms. Liquid marbles, the recently emerging microfluidic platform, could potentially resolve these drawbacks. This paper reports the first liquid marble based polymerase chain reaction. We demonstrated an experimental setup for the liquid-marble based PCR with a humidity-controlled chamber and an embedded thermal cycler. A concentrated salt solution was used to control the humidity of the PCR chamber which in turn reduces the evaporation rate of the liquid marble. The successful PCR of microbial source tracking markers for faecal contamination was achieved with the system, indicating potential application in water quality monitoring.

An automated on-demand liquid marble generator based on electrohydrodynamic pulling.

Liquid marble is a recently emerging digital microfluidic platform with a wide range of applications. Conventional liquid marbles are synthesized by coating liquid droplets with a thin layer of hydrophobic powder. Existing and emerging applications of liquid marbles require a contamination-free synthesis of liquid marbles with a high degree of reproducibility of their volume. Despite this requirement, the synthesis of liquid marbles has been still carried out manually. Manual production of liquid marbles leads to inconsistent volume and the possibility of contamination. The synthesis of liquid marbles with submicroliter volume is difficult to achieve and prone to large errors. This paper discusses the design and development of the first automated on-demand liquid marble generator with submicroliter capability. The device utilizes electrohydrodynamic pulling of liquid droplets on to a hydrophobic powder bed and subsequently coats them with the hydrophobic powder to synthesize liquid marbles of a desired volume.

Microfluidic Array Chip for Parallel Detection of Waterborne Bacteria.

The polymerase chain reaction (PCR) is a robust technique used to make multiple copies of a segment of DNA. However, the available PCR platforms require elaborate and time-consuming operations or costly instruments, hindering their application. Herein, we introduce a sandwiched glass-polydimethylsiloxane (PDMS)-glass microchip containing an array of reactors for the real-time PCR-based detection of multiple waterborne bacteria. The PCR solution was loaded into the array of reactors in a single step utilising capillary filling, eliminating the need for pumps, valves, and liquid handling instruments. Issues of generating and trapping bubbles during the loading chip step were addressed by creating smooth internal reactor surfaces. Triton X-100 was used to enhance PCR compatibility in the chip by minimising the nonspecific adsorption of enzymes. A custom-made real-time PCR instrument was also fabricated to provide thermal cycling to the array chip. The microfluidic device was successfully demonstrated for microbial faecal source tracking (MST) in water.

Manipulation of a floating liquid marble using dielectrophoresis.

A liquid marble is a microliter-sized droplet coated with hydrophobic powder. The porous coating prevents the liquid content from being in direct physical contact with its surroundings, making the liquid marble perfectly non-wetting. On the one hand, the non-wetting ability allows the liquid marble to float and move across a liquid surface with little resistance. On the other hand, the porosity enables gas exchange between the liquid marble and its surroundings. These properties allow the liquid marble to serve as a bioreactor platform for important applications such as cell culture. Liquid marbles floating on a free liquid surface prevent evaporation due to the high humidity near the liquid surface. Moving a floating liquid marble allows for stirring and mixing inside the liquid marble. This paper reports a novel technique for manipulating a floating liquid marble using dielectrophoresis. A relatively simple setup can move liquid marbles of various sizes across the water surface at high speeds. We also present an analytical model to model and accurately predict the motion of the floating liquid marble. The technique reported here potentially allows for high-throughput and efficient handling of floating liquid marbles as a digital microfluidics platform.

Digital polymerase chain reaction technology - recent advances and future perspectives.

Digital polymerase chain reaction (dPCR) technology has remained a "hot topic" in the last two decades due to its potential applications in cell biology, genetic engineering, and medical diagnostics. Various advanced techniques have been reported on sample dispersion, thermal cycling and output monitoring of digital PCR. However, a fully automated, low-cost and handheld digital PCR platform has not been reported in the literature. This paper attempts to critically evaluate the recent developments in techniques for sample dispersion, thermal cycling and output evaluation for dPCR. The techniques are discussed in terms of hardware simplicity, portability, cost-effectiveness and suitability for automation. The present paper also discusses the research gaps observed in each step of dPCR and concludes with possible improvements toward portable, low-cost and automatic digital PCR systems.

Cryoprotectant-Free Freezing of Cells Using Liquid Marbles Filled with Hydrogel.

Cryopreservation without cryoprotectant remains a significant challenge for the re-establishment of cell culture after freeze-thaw. Thus, finding an alternative and a simple cryopreservation method is necessary. Liquid marble (LM)-based digital microfluidics is a promising approach for cryoprotectant-free cryopreservation. However, the use of this platform to efficiently preserve samples with low cell density and well-controlled serum concentrations has not been investigated. We addressed this issue by embedding an agarose-containing fetal bovine serum (FBS) inside the LM. A low density of 500 cells/µL of murine 3T3 cells was selected for evaluating the postcryogenic survivability. The effects on the post-thaw cell viability of the concentration of agarose, the amount of FBS inside the agarose, and the volume of the LM were investigated systematically. This paper also presents an analysis on the changes in shape and crack size of post-thawed agarose. The results revealed that the embedded agarose gel serves as a controlled release mechanism of FBS and significantly improves cell viability. Post-thaw recovery sustains major cellular features, such as viability, cell adhesion, and morphology. The platform technology reported here opens up new possibilities to cryopreserve rare biological samples without the toxicity risk of cryoprotectants.

Evaporation dynamics of liquid marbles at elevated temperatures.

Study of evaporation dynamics of liquid marbles at elevated temperature is essential to determine the feasibility of liquid marbles to be used as micro compartments for digital polymerase chain reaction (PCR). We have modified an existing theoretical model of evaporation of a liquid droplet and verified its applicability on the evaporation of liquid marbles. The evaporation dynamics of an individual and a group of liquid marbles are analysed. This paper demonstrates that the evaporation dynamics of liquid marbles obeys the theoretical framework for elevated temperatures. The evaporation of a group of liquid marbles are observed as a coupled function of their diameter, their number in a group, the vapour density of the surrounding atmosphere and their spatial distribution.