Microfluidic Thermal Flowmeters for Drug Injection Monitoring.

This paper presents a microfluidic thermal flowmeter for monitoring injection pumps, which is essential to ensure proper patient treatment and reduce medication errors that can lead to severe injury or death. The standard gravimetric method for flow-rate monitoring requires a great deal of preparation and laboratory equipment and is impractical in clinics. Therefore, an alternative to the standard method suitable for remote, small-scale, and frequent infusion-pump monitoring is in great demand. Here, we propose a miniaturized thermal flowmeter consisting of a silicon substrate, a platinum heater layer on a silicon dioxide thin-membrane, and a polymer microchannel to provide accurate flow-rate measurement. The present thermal flowmeter is fabricated by the micromachining and micromolding process and exhibits sensitivity, linearity, and uncertainty of 0.722 mW/(g/h), 98.7%, and (2.36 ± 0.80)%, respectively, in the flow-rate range of 0.5-2.5 g/h when the flowmeter is operated in the constant temperature mode with the channel width of 0.5 mm. The measurement range of flow rate can be easily adjusted by changing the cross-sectional microchannel dimension. The present miniaturized thermal flowmeter shows a high potential for infusion-pump calibration in clinical settings.

Predictive performance of a new pharmacokinetic model for propofol in underweight patients during target-controlled infusion.

BACKGROUND: In a previous study, the modified Marsh and Schnider models respectively showed negatively- and positively-biased predictions in underweight patients. To overcome this drawback, we developed a new pharmacokinetic propofol model-the Choi model-for use in underweight patients. In the present study, we evaluated the predictive performance of the Choi model. METHODS: Twenty underweight patients undergoing elective surgery received propofol via TCI using the Choi model. The target effect-site concentrations (Ces) of propofol were 2.5, 3, 3.5, 4, 4.5, and 2 µg/mL. Arterial blood samples were obtained at least 10 minutes after achieving pseudo-steady-state. Predicted propofol concentrations with the modified Marsh, Schnider, and Eleveld pharmacokinetic models were obtained by simulation (Asan pump, version 2.1.3; Bionet Co. Ltd., Seoul, Korea). The predictive performance of each model was assessed by calculation of four parameters: inaccuracy, divergence, bias, and wobble. RESULTS: A total of 119 plasma samples were used to determine the predictive performance of the Choi model. Our evaluation showed that the pooled median (95% CI) bias and inaccuracy were 4.0 (-4.2 to 12.2) and 23.9 (17.6-30.3), respectively. The pooled biases and inaccuracies of the modified Marsh, Schnider, and Eleveld models were clinically acceptable. However, the modified Marsh and Eleveld models consistently produced negatively biased predictions in underweight patients. In particular, the Schnider model showed greater inaccuracy at a target Ce ≥ 3 µg/mL. CONCLUSION: The new propofol pharmacokinetic model (the Choi model) developed for underweight patient showed adequate performance for clinical use.

Portable Skin Analyzers with Simultaneous Measurements of Transepidermal Water Loss, Skin Conductance and Skin Hardness.

Simultaneous measurement of skin physiological and physical properties are important for the diagnosis of skin diseases and monitoring of human performance, since it provides more comprehensive understanding on the skin conditions. Current skin analysis devices, however, require each of probes and unique protocols for the measurement of individual skin properties, resulting in inconvenience and increase of measurement uncertainty. This paper presents a pen-type skin analyzing device capable tomeasure three key skin properties at the same time: transepidermal water loss (TEWL), skin conductance, and skin hardness. It uses a single truncated hollow cone (THC) probe integrated with a humidity sensor, paired electrodes, and a load cell for the multimodal assessment of the skin properties. The present device measured TEWL with a sensitivity of 0.0068 (%/s)/(g/m2/h) and a linearity of 99.63%, conductance with a sensitivity of 1.02 µS/µS and a linearity of 99.36%, and hardness with a sensitivity of 0.98 Shore 00/Shore 00 and a linearity of 99.85%, within the appropriate ranges for the human skin. The present pen-type device has a high potential for the skin health diagnosis as well as the human performance monitoring applications.

A High Throughput Apoptosis Assay using 3D Cultured Cells.

A high throughput apoptosis assay using 3D cultured cells was developed with a micropillar/microwell chip platform. Live cell apoptosis assays based on fluorescence detection have been useful in high content screening. To check the autofluorescence of drugs, controls (no caspase-3/7 reagent in the assay) for the drugs are necessary which require twice the test space. Thus, a high throughput capability and highly miniaturized format for reducing reagent usage are necessary in live cell apoptosis assays. Especially, the expensive caspase-3/7 reagent should be reduced in a high throughput screening system. To solve this issue, we developed a miniaturized apoptosis assay using micropillar/microwell chips for which we tested seventy drugs (six replicates) per chip and reduced the assay volume to 1 µL. This reduced assay volume can decrease the assay costs compared to the 10-40 µL assay volumes used in 384 well plates. In our experiments, among the seventy drugs, four drugs (Cediranib, Cabozatinib, Panobinostat, and Carfilzomib) induced cell death by apoptosis. Those results were confirmed with western blot assays and proved that the chip platform could be used to identify high potency apoptosis-inducing drugs in 3D cultured cells with alginate.

3D Cell-Based High-Content Screening (HCS) Using a Micropillar and Microwell Chip Platform.

A micropillar/microwell chip platform was applied to develop a 3D cell-based high-content screening (HCS) platform. Previously, 3D cell culture in the micropillar/microwell chip platform was only limited to cell viability measurements in a high-throughput manner. However, an HCS system could provide biological and phenotypic information which was very useful to understand complex biological functions and mechanisms of drug actions. To stain 3D cultured cells immobilized with alginate spots, we developed and optimized antibody staining procedures for 3D cultured cells. As a proof of concept, the phospho-EGFR (p-EGFR, a highly expressed receptor protein in cancer), F-actin (a protein of the actin cytoskeleton), and nuclei of 3D cultured cells were stained and analyzed after being treated with 72 different drugs. The p-EGFR inhibition of the drugs was successfully identified in the 3D cultured cells by comparing p-EGFR expression with that of F-actin and the nucleus. The p-EGFR expression levels were also measured by Western blot to verify the chip data.

Tapered-slit membrane filters for high-throughput viable circulating tumor cell isolation.

This paper presents tapered-slit membrane filters for high-throughput viable circulating tumor cell (CTC) isolation. The membrane filter with a 2D array of vertical tapered slits with a gap that is wide at the entrance and gradually decreases with depth, provide minimal cell stress and reduce 82.14% of the stress generated in conventional straight-hole filters. We designed two types of tapered-slit filters, Filters 6 and 8, respectively, containing the tapered slits with outlet widths of 6 µm and 8 µm at a slit density of 34,445/cm(2) on the membrane. We fabricated the vertical slits with a tapered angle of 2 ° on a SU8 membrane by adjusting the UV expose dose and the air gap between the membrane and the photomask during lithography. In the experimental study, the proposed tapered-slit filter captured 89.87% and 82.44% of the cancer cells spiked in phosphate buffered saline (PBS) and diluted blood (blood: PBS = 1:4), respectively, at a sample flow rate of 5 ml per hour, which is 33.3 times faster than previous lateral tapered-slit filters. We further verified the capability to culture on chip after capturing: 72.33% of cells among the captured cells still remained viable after a 5-day culture. The proposed tapered-slit membrane filters verified high-throughput viable CTC isolation capability, thereby inaugurating further advanced CTC research for cancer diagnosis and prognosis.

Sub-population analysis of deformability distribution in heterogeneous red blood cell population.

We present a method for sub-population analysis of deformability distribution using single-cell microchamber array (SiCMA) technology. It is a unique method allowing the correlation of overall cellular characteristics with surface and cytosolic characteristics to define the distribution of individual cellular characteristics in heterogeneous cell populations. As a proof of principle, reticulocytes, the immature sub-population of red blood cells (RBC), were recognized from RBC population by a surface marker and different characteristics on deformability between these populations were characterized. The proposed technology can be used in a variety of applications that would benefit from the ability to measure the distribution of cellular characteristics in complex populations, especially important to define hematologic disorders.

Clinical Validation of a Wearable Piezoelectric Blood-Pressure Sensor for Continuous Health Monitoring.

Wearable blood-pressure sensors have recently attracted attention as healthcare devices for continuous non-invasive arterial pressure (CNAP) monitoring. However, the accuracy of wearable blood-pressure (BP) monitoring devices has been controversial due to the low signal quality of sensors, the absence of an accurate transfer function to convert the sensor signals into BP values, and the lack of clinical validation regarding measurement precision. Here, a wearable piezoelectric blood-pressure sensor (WPBPS) is reported, which achieves a high normalized sensitivity (0.062 kPa-1 ), and fast response time (23 ms) for CNAP monitoring. The transfer function of a linear regression model is designed, offering a simple solution to convert the flexible piezoelectric sensor signals into BP values. In order to verify the measurement accuracy of WPBPS, clinical trials are performed on 35 subjects aged from 20 to 80 s after screening. The mean difference between the WPBPS and a commercial sphygmomanometer of 175 BP data pairs is -0.89 ± 6.19 and -0.32 ± 5.28 mmHg for systolic blood pressure (SBP) and diastolic blood pressure (DBP), respectively. By building a WPBPS-embedded wristwatch, the potentially promising use of a convenient, portable, continuous BP monitoring system for cardiovascular disease diagnosis is demonstrated.

Deep learning-based robust automatic non-invasive measurement of blood pressure using Korotkoff sounds.

This paper proposes a method that automatically measures non-invasive blood pressure (BP) based on an auscultatory approach using Korotkoff sounds (K-sounds). There have been methods utilizing K-sounds that were more accurate in general than those using cuff pressure signals only under well-controlled environments, but most were vulnerable to the measurement conditions and to external noise because blood pressure is simply determined based on threshold values in the sound signal. The proposed method enables robust and precise BP measurements by evaluating the probability that each sound pulse is an audible K-sound based on a deep learning using a convolutional neural network (CNN). Instead of classifying sound pulses into two categories, audible K-sounds and others, the proposed CNN model outputs probability values. These values in a Korotkoff cycle are arranged in time order, and the blood pressure is determined. The proposed method was tested with a dataset acquired in practice that occasionally contains considerable noise, which can degrade the performance of the threshold-based methods. The results demonstrate that the proposed method outperforms a previously reported CNN-based classification method using K-sounds. With larger amounts of various types of data, the proposed method can potentially achieve more precise and robust results.

Multi-volume hemacytometer.

Cell counting has become an essential method for monitoring the viability and proliferation of cells. A hemacytometer is the standard device used to measure cell numbers in most laboratories which are typically automated to increase throughput. The principle of both manual and automated hemacytometers is to calculate cell numbers with a fixed volume within a set measurement range (105 ~ 106 cells/ml). If the cell concentration of the unknown sample is outside the range of the hemacytometer, the sample must be prepared again by increasing or decreasing the cell concentration. We have developed a new hemacytometer that has a multi-volume chamber with 4 different depths containing different volumes (0.1, 0.2, 0.4, 0.8 µl respectively). A multi-volume hemacytometer can measure cell concentration with a maximum of 106 cells/ml to a minimum of 5 × 103 cells/ml. Compared to a typical hemacytometer with a fixed volume of 0.1 µl, the minimum measurable cell concentration of 5 × 103 cells/ml on the multi-volume hemacytometer is twenty times lower. Additionally, the Multi-Volume Cell Counting model (cell concentration calculation with the slope value of cell number in multi-chambers) showed a wide measurement range (5 × 103 ~ 1 × 106 cells/ml) while reducing total cell counting numbers by 62.5% compared to a large volume (0.8 µl-chamber) hemacytometer.

Passive flow-rate regulators using pressure-dependent autonomous deflection of parallel membrane valves.

We present passive flow-rate regulators using an autonomous deflection of parallel membrane valves, capable to maintain a constant flow-rate at varying inlet pressure supplied from micropumps. The previous passive flow-rate regulators are difficult to integrate with micropumps, not only because of the complex multi-layer structures, but also because of the high threshold inlet pressure required for flow-rate regulation. In this study, we present passive flow-rate regulators using parallel membrane valves, capable of achieving flow-rate regulation function at the minimum threshold inlet pressure as low as 15 kPa with simple structure formed by a single mask process. The parallel membranes in a flow-rate regulator are designed to deflect and adjust flow resistance autonomously according to the inlet pressure, thus maintaining a constant flow-rate independent of the inlet pressure variation. We designed the four different prototypes of W20, W30, W40, and W50, having parallel membrane widths of 20, 30, 40 and 50 microm, respectively. We estimated the flow-rate based on both analytical and numerical models. In an experimental study, we observed the deformation of parallel membranes and the flow-rate depending on the inlet pressure. The fabricated prototypes achieved the constant flow-rate of 6.09 +/- 0.32 microl s(-1) (W20 fabricated by 10 : 1 PDMS (PolyDiMethylSiloxane)) over an inlet pressure of 20 kPa. We also observed that prototypes fabricated by 20 : 1 PDMS, having lower Young's modulus than normal 10 : 1 PDMS, showed a lower threshold pressure and higher regulated flow-rate than prototypes fabricated by 10 : 1 PDMS. W40 fabricated by 20 : 1 PDMS showed a constant flow-rate of 14.53 +/- 0.51 microl s(-1) over inlet pressure of 15 kPa. The present passive flow-rate regulators have strong potential for applications in integrated microfluidic systems.

A three-dimensional (3D) particle focusing channel using the positive dielectrophoresis (pDEP) guided by a dielectric structure between two planar electrodes.

We present a three-dimensional (3D) particle focusing channel using the positive dielectrophoresis (pDEP) guided by a dielectric structure between two planar electrodes. The dielectric structure between two planar electrodes induces the maximum electric field at the center of the microchannel and particles are focused to the center of the microchannel by pDEP as they flow from the single sample injection port. Compared to the previous 3D particle focusing methods using standing surface acoustic wave (SSAW), hydrodynamic force, and negative dielectrophoresis (nDEP), the present device achieves the simple and effective particle focusing function without any additional fluidic ports and top electrodes. The present focusing channel is also fabricated by PDMS and glass substrate with electrodes, compatible for the integrated microbiochemical analysis system. We designed and fabricated the particle focusing channel based on the numerical estimation of particle position and focusing efficiency. In the experimental study, approximately 90% focusing efficiency was achieved within the focusing length of 2 mm, on both the x-z plane (top-view) and y-z plane (side-view) for 2 microm-diameter polystyrene (PS) beads at the applied voltage over 15 V(p-p) (square wave) and at a flow rate below 0.01 microl/min. Focusing experiments using 4.5 microm-diameter PS beads and yeast cells also verified that the present focusing channel can be applied to bio-particles having different sizes and properties. The present simple 3D particle focusing channel is suitable for use in integrated microbiochemical analysis systems.

Drug Efficacy Comparison of 3D Forming and Preforming Sphere Models with a Micropillar and Microwell Chip Platform.

Hepatocellular carcinoma (HCC), a major histological subtype of liver cancer, is the third most common cause of cancer-related death worldwide. Currently, many curative standard treatments using target-specific chemotherapeutic agents are being developed. However, drug efficacy tests based on the 2D monolayer cell culture model do not effectively screen the best drug candidates because they do not accurately reflect in vivo tumor microenvironments. Thus, to select the best drug candidates or repositioning drugs, we developed new 3D in vitro hepatic tumor models, including 3D forming and preformed sphere models. A micropillar and microwell chip platform was used for the 3D in vitro liver cell-based model for high-throughput screening. We measured the efficacy of 60 drugs and sorted the most efficacious drugs by comparing the drug response of the 2D monolayer model with the 3D forming and preformed sphere models. Among the 60 drugs, 17 drugs (28.3%) showed a significant high efficacy in the 3D preformed sphere model, while 45 drugs (75%) showed an efficacy in the 2D model. We also calculated the IC50 values of the 17 drugs and found that 7 drugs exhibited a high sensitivity in HCC, which was in agreement with previous studies.

Retina phantom for the evaluation of optical coherence tomography angiography based on microfluidic channels.

Optical coherence tomography (OCT) angiography (OCTA) has been actively studied as a noninvasive imaging technology to generate retinal blood vessel network maps for the diagnoses of retinal diseases. Given that the uses of OCT and OCTA have increased in the field of ophthalmology, it is necessary to develop retinal phantoms for clinical OCT for product development, performance evaluation, calibration, certification, medical device licensing, and production processes. We developed a retinal layer-mimicking phantom with microfluidic channels based on microfluidic fabrication technology using polydimethylsiloxane (PDMS) and titanium dioxide (TiO2) powder. We implemented superficial and deep retinal vessels using microfluidic channels. In addition, multilayered thin films were synthesized with multiple spin-coating processes that comprised layers that corresponded to the retinal layers, including the ganglion cell layer (GCL), inner plexiform layer (IPL), and inner nuclear layer (INL). The phantom was formed by merging the multilayered thin film, and microfluidic channels were assembled with an optical lens, water chamber, and an aluminum tube case. Finally, we obtained cross-sectional OCT images and en-face OCTA images of the retinal phantom using lab-made ophthalmic OCT. From the cross-sectional OCT image, we could compare each of the layer thicknesses of the phantom with the corresponding layer thicknesses of the human retina. In addition, we obtained en-face OCTA images with injections of intralipid solutions. It is shown that this phantom will be able to be potentially used as a convenient tool to evaluate and standardize the quality and accuracy of OCT and OCTA images.

Accuracy assessment of a PION TCI pump based on international standards.

BACKGROUND: Inaccuracies associated with target-controlled infusion (TCI) delivery systems are attributable to both software and hardware issues, as well as pharmacokinetic variability. However, little is known about the inaccuracy of the syringe pump operating in TCI mode. This study aimed to evaluate the accuracy of the TCI pump based on international standards. METHODS: A test apparatus for accuracy evaluation of a syringe pump (PION TCI®, Bionet Co. Ltd.) was designed to apply the gravimetric method. Pump accuracy was evaluated in terms of deviation defined by the following equation: infusion rate deviation (%) = (Ratemea - Rateest ) / Rateest × 100, where Ratemea is the infusion rate (ml/h) as measured by the gravimetric system, and Rateest is the infusion rate (ml/h) as estimated by the pump. An infusion rate representing TCI mode was determined from previous clinical trial data which evaluated the predictive performance of the pharmacokinetic model. The PION TCI pump used in that clinical trial was used to evaluate accuracy of the syringe pump. The distribution of infusion rates obtained from the clinical trial was calculated, and the median value of the distribution was determined as the representative value. RESULTS: The representative infusion rate representing TCI mode was 31 ml/h, at which the infusion rate deviation was 4.5 ± 1.6%. CONCLUSIONS: The inaccuracy of the syringe pump contributing to TCI system inaccuracy is insignificant.

Truncated Hollow Cone Probe for Assessing Transepidermal Water Loss and Skin Hardness.

This study proposes a skin analysis device using a truncate hollow cone (THC) probe for measuring both transepidermal water loss (TEWL) and skin hardness. Because skin health is closely related to the epidermal barrier function and skin mechanical property, it is important to measure their biophysical indicators at the same time, to understand skin conditions and diagnose skin disorders such as atopic dermatitis and systemic sclerosis. Previous skin analyzers, however, required different probes with different protocols for each biophysical indicators, which makes the measurement inconvenient and increases the measurement uncertainty. The present device consists of a THC probe equipped with humidity and force sensors, and an actuator that simultaneously measure both TEWL and skin hardness which indicate the integrity of the epidermal barrier function and the skin mechanical property, respectively. Using artificial reference skins, the prototype device showed the TEWL with a sensitivity and linearity of 0.011 (%/s)/(g/m2/h) and 99.5%, and the hardness with 0.075 N/(Shore 00) and 97.6%, respectively, which are within the appropriate range for the properties of human skin. The on-body measurement of five subjects showed that the proposed device could measure both the TEWL and skin hardness without any crosstalk from each other. The proposed device has great potential for in-depth analysis of the health status of the skin which could indicate various skin diseases.

Realization of an ultrathin acoustic lens for subwavelength focusing in the megasonic range.

In this study, we report the first experimental realization of an ultrathin (0.14λ, λ = 1.482 mm means wavelength at 1 MHz in the water medium) subwavelength focusing acoustic lens that can surpass the Rayleigh diffraction limit (0.61λ/NA, NA means numerical aperture). It is termed a Super-Oscillatory Acoustic Lens (SOAL), and it operates in the megasonic range. The SOAL represents an interesting feature allowing the achievement of subwavelength focusing without the need to operate in close proximity to the object to be imaged. The optimal layout of the SOAL is obtained by utilizing a systematic design approach, referred to here as topology optimization. To this end, the optimization formulation is newly defined. The optimized SOAL is fabricated using a photo-etching process and its subwavelength focusing performance is verified experimentally via an acoustic intensity measurement system. From these measurements, we found that the proposed optimized SOAL can achieve superior focusing features with a Full Width at Half Maximum (FWHM) of ~0.40λ/NA ≃ 0.84 mm (for our SOAL, NA = 0.707) with the transmission efficiency of 26.5%.

Volumetric Analysis of 3-D-Cultured Colonies in Wet Alginate Spots Using 384-Pillar Plate.

The volumetric analysis of three-dimensional (3-D)-cultured colonies in alginate spots has been proposed to increase drug efficacy. In a previously developed pillar/well chip platform, colonies within spots are usually stained and dried for analysis of cell viability using two-dimensional (2-D) fluorescent images. Since the number of viable cells in colonies is directly related to colony volume, we proposed the 3-D analysis of colonies for high-accuracy cell viability calculation. The spots were immersed in buffer, and the 3-D volume of each colony was calculated from the 2-D stacking fluorescent images of the spot with different focal positions. In the experiments with human gastric carcinoma cells and anticancer drugs, we compared cell viability values calculated using the 2-D area and 3-D volume of colonies in the wet and dried alginate spots, respectively. The IC50 value calculated using the 3-D volume of the colonies (9.5 µM) was less than that calculated in the 2-D area analysis (121.5 µM). We observed that the colony showed a more sensitive drug response regarding volume calculated from the 3-D image reconstructed using several confocal images than regarding colony area calculated in the 2-D analysis.

Label-free Rapid Viable Enrichment of Circulating Tumor Cell by Photosensitive Polymer-based Microfilter Device.

We present a clinical device for simple, rapid, and viable isolation of circulating tumor cells (CTCs) from cancer patient bloods. In spite of the clinical importance of CTCs, the lack of easy and non-biased isolation methods is a big hurdle for implementing CTC into clinical use. The present device made of photosensitive polymer was designed to attach to conventional syringe to isolate the CTCs at minimal resources. Its unique tapered-slits on the filter are capable not only to isolate the cell based on their size and deformability, but also to increase sample flow rate, thus achieving label-free rapid viable CTC isolation. We verified our device performance using 9 different types of cancer cells at the cell concentration from 5 to 100cells/ml, showing that the device capture 77.7% of the CTCs while maintaining their viability of 80.6%. We extended our study using the 18 blood samples from lung, colorectal, pancreatic and renal cancer patients and captured 1-172 CTCs or clustered CTCs by immunofluorescent or immunohistochemical staining. The captured CTCs were also molecularly assayed by RT-PCR with three cancer-associated genes (CK19, EpCAM, and MUC1). Those comprehensive studies proved to use our device for cancer study, thereby inaugurating further in-depth CTC-based clinical researches.

High-Dose Compound Heat Map for 3D-Cultured Glioblastoma Multiforme Cells in a Micropillar and Microwell Chip Platform.

Glioblastoma multiforme (GBM) is recognized as the most common and lethal form of central nervous system cancer. To cure GBM patients, many target-specific chemotherapeutic agents have been developing. However, 2D monolayer cell-based toxicity and efficacy tests did not efficiently screen agents due to the pool reflection of in vivo microenvironments (cell-to-cell and cell-to-extracellular matrix interaction). In this study, we used a 3D cell-based, high-throughput screening method reflecting the microenvironments using a micropillar and microwell chip platform to draw a high-dose heat map of the cytotoxicity and efficacy of 70 compounds, with two DMSO controls. Moreover, the high-dose heat map model compared the responses of four 3D-cultured patient-derived GBM cells and astrocytes to high dosages of compounds with respect to efficacy and cytotoxicity, respectively, to discern the most efficacious drug for GBM. Among the 70 compounds tested, cediranib (a potent inhibitor of vascular endothelial growth factor (VEGF) receptor tyrosine kinases) exhibited the lowest cytotoxicity to astrocytes and high efficacy to GBM cells in a high-dose heat map model.