This work was sponsored by the Ministry of Science and Technology, Taiwan, under grant numbers MOST 104-2221-E-241-001-MY3 and MOST 105-2627-B-005-002.

1. Experimental Chip Module
<ol>
	<li>Fabricate the indium tin oxide (ITO) electrode chip by photolithography and chemical wet etching processes
	<ol>
		<li>Obtain an ITO substrate (370 mm x 480 mm x 0.5 mm (L x W x H)) with a 2,600 &#197; ITO layer commercially (See Materials List). Slice the ITO substrate to the dimensions of 90 mm x 90 mm x 0.5 mm with a glass cutter for the ITO electrode patterning process in a 4 inch aligner.</li>
		<li>Use an ultrasonic cleaner to clean the ITO glass with acetone and then with deionized water, for 15 min each. Dry the ITO glass with clean dry air.</li>
		<li>Dispense 5 ml of positive photoresist solution onto the surface of the ITO glass.</li>
		<li>Use spin coater at 500 x g for 30 sec to produce a uniform photoresist layer. Then bake on a hotplate at 90 &#176;C for 5 min to drive off excess solvent in the photoresist.</li>
		<li>Expose the ITO glass to 14 mW of ultraviolet light at 436 nm for 3.1 sec through a film photomask with the designed pattern (See Materials List).</li>
		<li>Immerse the sample in 60 ml development solution at 23 &#176;C for 30 sec to develop the patterned routes. Then bake on a hotplate at 120 &#176;C for 10 min to harden the photoresist and improve photoresist adhesion.</li>
		<li>Immerse the sample for 3 min in 60 ml etching solution at 80 &#176;C to etch the unprotected ITO layer.</li>
		<li>Immerse the sample for 1 min in 60 ml acetone to remove photoresist on surface of the ITO glass.</li>
		<li>Slice the ITO glass into the dimensions of 62 mm x 35 mm for the experimental ITO electrode chip (Figure 1) with a glass cutter.</li>
	</ol>
	</li>
</ol>
<img alt="Figure 1" src="/files/ftp_upload/54575/54575fig1.jpg" /> 
Figure 1: ITO electrode chip. The fabricated ITO chip with 8 pairs of electrode-patterned routes is shown. There are 15 electrodes measuring 2 mm x 8 mm at the side edge, and the central two routes share the same electrode. The distance between each pair of electrode fingers in a test well is 7 mm. <a href="https://www-jove-com.remotexs.ntu.edu.sg/files/ftp_upload/54575/54575fig1large.jpg" target="_blank">Please click here to view a larger version of this figure.</a>
<ol start="2">
	<li>Construct the experimental chip module
	<ol>
		<li>Clean the commercial 8-well silicone array with an ultrasonic cleaner as shown in Figure 2 with detergent, then deionized water, then 95% ethanol, and then deionized water, for 15 min each.</li>
		<li>Dry the 8-well silicone array by blowing clean dry air.</li>
		<li>Press the 8-well silicone array into the ITO chip in order to form the experimental chip module (Figure 3). Tightly bind the silicone array and ITO chip.</li>
	</ol>
	</li>
</ol>
<img alt="Figure 2" src="/files/ftp_upload/54575/54575fig2.jpg" /> 
Figure 2: Silicone well array. The commercial 8-well silicone array can hold 8 tested samples simultaneously. The size of each well is 11 mm x 8 mm x 8.5 mm (L x W x H). <a href="https://www-jove-com.remotexs.ntu.edu.sg/files/ftp_upload/54575/54575fig2large.jpg" target="_blank">Please click here to view a larger version of this figure.</a>
<img alt="Figure 3" src="/files/ftp_upload/54575/54575fig3.jpg" /> 
Figure 3: Experimental chip module. The ITO electrode chip is attached with the 8-well silicone array to form the experimental chip module. The adhesion between the silicone array and the ITO chip is strong. Therefore, the silicone array and the ITO chip can bond together for use without any adhesive substance. <a href="https://www-jove-com.remotexs.ntu.edu.sg/files/ftp_upload/54575/54575fig3large.jpg" target="_blank">Please click here to view a larger version of this figure.</a>
2. Impedance Measurement
<ol>
	<li>Connect the personal computer, lock-in amplifier, and switch relay to form the impedance readout module as shown in Figure 4.</li>
</ol>
<img alt="Figure 4" src="/files/ftp_upload/54575/54575fig4.jpg" /> 
Figure 4: Schematic of the impedance-based apparatus. The lock-in amplifier, switch relay, and personal computer comprise the impedance readout module. The commercial phase-sensitive lock-in amplifier is used to send and extract the electrical signals. The homemade switch relay circuit connecting various ITO chips is used to specify which well and which ITO chip to be tested. A total of 6 chips can be connected to the switch relay specifying 48 samples in a time sharing manner. The real-time in-phase resistance and the signal phase shift of the tested solution are recorded continuously on a personal computer for the whole evaporation process. <a href="https://www-jove-com.remotexs.ntu.edu.sg/files/ftp_upload/54575/54575fig4large.jpg" target="_blank">Please click here to view a larger version of this figure.</a>
<ol start="2">
	<li>Put the experimental chip module into the socket of the switch relay.</li>
	<li>Input parameters in the computer program. Input the signal frequency (1 kHz), the specified well number (0-7), the execution cycle (100), and the filename (HA).</li>
</ol>
3. Evaporation Experiments
<ol>
	<li>Prepare four 2.5 ml hyaluronic acid solutions at 0, 0.05, 0.5 and 1 w/v% in water. Place each 2.5 ml sample solution in a vial measuring 14.75 mm x 45 mm x 8 mm (O.D. x H x I.D.).</li>
	<li>For each solution, add 0.5 ml sample solution to a single well of the ITO chip module.</li>
	<li>Weigh and record the initial weight of each vial by the electronic balance machine.</li>
	<li>Execute the computer program to automatically measure and record the real-time in-phase resistance and the signal phase shift of specified wells on the ITO chip.</li>
	<li>Start the evaporation experiments simultaneously in the same place by both the weighing method and impedance method.</li>
	<li>Weigh and record the weight of each vial by the electronic balance machine at scheduled time points.</li>
	<li>Analyze collected data in the weighing method and impedance method.19</li>
</ol>

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The authors have nothing to disclose.

The critical step for evaporation measurement in this impedance-based detection is the preparation of the tested solutions. Deionized water cannot be used due to its enormous impedance. Instead, tap water containing conductive ions was used to prepare hyaluronic acid solutions for experiments. However, the electrical properties of tap water were not constant for use. Therefore, normalization, such as the relative evaporation rate to water in this study, was adopted as an alternative index for evaporation. The limitation of this technique is that tested solutions must have conductive ions for electrochemical characterization.
Very recently, a graphene-based impedance chip has been proposed for the modification of this technique20. With exceptional electronic and optoelectronic properties, graphene has attained considerable attention as an alternative to ITO for various electrode or conductor applications. The graphene-based finger-like electrode chip was successfully demonstrated in examining the stability of emulsion products by electrochemical impedance spectroscopy.
This study revealed that a 0.05% hyaluronic acid solution can reduce the relative evaporation rate to water by 12% as measured by the impedance. Therefore, topical application of 0.1% hyaluronic acid cream can lead to a significant improvement in skin hydration21. The molecular weight of hyaluronic acid plays an important role in its applications. For example, hyaluronic acid with a higher molecular weight could have better analgesic effects22. The application of low-molecular-weight hyaluronic acid had a significant reduction of wrinkle depth due to better penetration abilities21. In the future, the effects of the molecular weight on the moisturizing capacity of hyaluronic acid can be studied simultaneously on this impedance-based platform with multiple sample measurements for comparison purposes. A total of 6 chips can be connected to the homemade switch relay specifying the well to be tested for a real time test on 48 samples in a time sharing manner.
Although the conventional weight change approach stands as a simple and the most straightforward way to measure the moisturizing capacity of a solution, it is a time-consuming approach for observing enough weight change to determine an accurate evaporation rate. For example, it took about half a day to detect the desired evaporation rate of hyaluronic acid solution due to the detection limit of a precision balance with reasonable experimental error in this study. However, the electrical property of a solution is more sensitive than weight. The change in electrical properties can be detected sooner than weight loss in the evaporation process. In this study, the change rate in electrical impedance of hyaluronic acid solution at the end of a one hour observation period of evaporation was sufficiently determined. Therefore, the presented impedance-based detection apparatus is found to outperform the conventional weighing method in terms of detection sensitivity and response time.
Corresponding to the previous publication23 and commercial device for the assessment of transdermal water loss, the electrical property can be treated as an index to reflect the evaporation rate. However, this presented impedance-based detection apparatus shows following advantages over the former: (i) a small sample volume requirement, (ii) parallel detection, (iii) easy disassembly for cleaning and reuse, and (iv) multiple applications such as bio molecular detection, cellular behavior, and phase separation16-19. The proposed high-sensitivity and fast-response impedance-based apparatus is validated as a superior candidate to handle evaporation tests relative to a conventional weight loss approach. In the future, this proposal impedance-based apparatus can also potentially be applied in any intrinsic property of a material or a specific process that could affect the conductivity of an electrochemical system24.

Evaporation is a type of liquid vaporization and occurs along the gas-liquid interface of a collective body of water. The water molecules near the surface become capable of escaping from the liquid due to collision of water molecules. The evaporation rate is an important key factor during the process of evaporation. Generally, a balance or volumetric tube1-3 is widely-used to detect the evaporation of solutions. However, it takes a long time to measure the evaporation rate due to the precision limitation of a balance or a volumetric tube. For this reason, a responsive and high-sensitivity instrument must be developed to probe into the details of the evaporation process.
Electrochemical impedance spectroscopy (EIS) is a fast-response, sensitive and effective experimental means in terms of in-situ impedance detection for electrochemical system characterization4. Therefore, EIS can be applied in various fields, such as recent studies on cellular behavior5, bioanalytical sensing6-7, electrolysis8, conducting polymers9, and electrochemical extraction10. Even though EIS systems had successfully been applied in a wide variety of disciplines, there exist an extremely small number of publications on its application to evaporation research.
Hyaluronic acid, a high molecular weight polysaccharide with strong water-binding potential, is a well-known humectant for cosmetic applications. One hyaluronic acid molecule can bind up to 500 water molecules11 and reach 1,000 times its original volume12. An extremely small amount of hyaluronic acid can possess moisturizing function13-14. Due to the high moisture retention, hyaluronic acid has become an important component of cosmetic humectant products with high commercial value worldwide15.
This study presents the method of a novel impedance-based apparatus featuring high speed detection, small volume sample requirement, and multiple sample measurements16-19. It is presented with a focus on the relative evaporation rate comparison among solutions as a way to validate the superiority of the innovative detection mechanism over a conventional weighing manner.

<table><tbody><tr><td>95% ethanol</td><td>Echo Chemical Co., Ltd., Miaoli, Taiwan</td><td>484000001103C-00EC</td><td></td></tr><tr><td>Acetone</td><td>Avantor Performance Materials Inc., Center Valley, PA, USA</td><td>JTB-9005-68</td><td></td></tr><tr><td>Development solution</td><td>Kemitek Industrial Crop., Hsinchu, Taiwan</td><td>12F01031</td><td>KTD-1</td></tr><tr><td>Etching solution</td><td>eSolv Technology Co., Taipei, Taiwan</td><td></td><td>EG-462</td></tr><tr><td>Hyaluronic acid</td><td>Shandong Freda Biopharm Co., Ltd., Jinan, China</td><td>1010212</td><td>Molecular weight 980k, Cosmetic Grade</td></tr><tr><td>Photoresist solution</td><td>AZ Electronic Materials Taiwan Co., Ltd., Hsinchu, Taiwan</td><td>65101M19</td><td>AZ6112</td></tr><tr><td>8-well silicone array</td><td>Greiner bio-one Inc., Frickenhausen, Baden-W&amp;uuml;rttemberg, Germany</td><td>FlexiPERM</td><td></td></tr><tr><td>ITO glass</td><td>GemTech Optoelectronics Co., Taoyuan, Taiwan</td><td></td><td></td></tr><tr><td>Vial</td><td>Sigma-Aldrich Co. LLC., St. Louis, MO, USA</td><td>854190</td><td></td></tr><tr><td>Film photomask</td><td>Taiwan Mesh Co., Ltd, Taoyuan, Taiwan</td><td></td><td></td></tr><tr><td>Lock-in amplifier</td><td>Stanford Research Systems, Inc., Palo Alto, CA, USA</td><td>SR830</td><td></td></tr><tr><td>Switch relay</td><td>Instrument Technology Research Center, National Applied Research Laboratories, Hsinchu, Taiwan</td><td></td><td></td></tr><tr><td>Electronic balance machine</td><td>Radwag Inc., Radom, Poland</td><td>AS 60/220/C/2</td><td></td></tr></tbody></table>

a high performance impedance-based platform for evaporation rate detection

This paper describes the method of a novel impedance-based platform for the detection of the evaporation rate. The model compound hyaluronic acid was employed here for demonstration purposes. Multiple evaporation tests on the model compound as a humectant with various concentrations in solutions were conducted for comparison purposes. A conventional weight loss approach is known as the most straightforward, but time-consuming, measurement technique for evaporation rate detection. Yet, a clear disadvantage is that a large volume of sample is required and multiple sample tests cannot be conducted at the same time. For the first time in literature, an electrical impedance sensing chip is successfully applied to a real-time evaporation investigation in a time sharing, continuous and automatic manner. Moreover, as little as 0.5 ml of test samples is required in this impedance-based apparatus, and a large impedance variation is demonstrated among various dilute solutions. The proposed high-sensitivity and fast-response impedance sensing system is found to outperform a conventional weight loss approach in terms of evaporation rate detection.

During the evaporation process, the conductive ions in the tested solution became concentrated with the decreasing solution volume, and the impedance of this solution decreased. The rates of weight loss and impedance decrease in the evaporation progress for each tested solution were measured. For comparison purposes, the data in the rates of weight loss and impedance decrease were normalized to water and then plotted together in Figure 5. As illustrated in Figure 5, the weight loss demonstrates the same tendency as impedance, and shows that the relative evaporation rate to water evaporation decreases with the hyaluronic acid concentration. However, a large amount of variation is found in the proposed impedance-based approach than in the conventional weighing method for the evaporation examinations. The normalized data only had a 0.06 fall from 0% to 1% hyaluronic acid concentration intervention in the weighing approach, while a tremendous drop of 0.84 was found in the impedance-based apparatus. The simple linear equation is used to relate the normalized rates of weight loss and impedance decrease.
Y = 0.0852X + 0.9166, R2 = 0.97
where X and Y represent the normalized rates of impedance decrease and weight loss, respectively. The rate of weight loss, i.e., the evaporation rate of interest, in hyaluronic acid solution can be found correspondingly by way of the measured data in the impedance decrease. In practical applications, the measured impedance data can be quickly converted into the weight loss of hyaluronic acid solution by this linear equation.
<img alt="Figure 5" src="/files/ftp_upload/54575/54575fig5.jpg" /> 
Figure 5: Relative evaporation rates to water of hyaluronic acid solutions at different concentrations. The relative evaporation rate to water is defined as the evaporation rate of a solution normalized by water. The relative evaporation rate to water against hyaluronic acid concentration by tests of balance and impedance chip are shown together for comparison. There is a larger change in the testing of impedance chip as compared the testing of balance. The error bar is the standard deviation in three experiments. <a href="https://www-jove-com.remotexs.ntu.edu.sg/files/ftp_upload/54575/54575fig5large.jpg" target="_blank">Please click here to view a larger version of this figure.</a>

Watch this Scientific Journal Video about A High Performance Impedance-based Platform for Evaporation Rate Detection at JoVE.com

A High Performance Impedance-based Platform for Evaporation Rate Detection

This paper presents an impedance-based apparatus for evaporation rate detection of solutions. It offers clear advantages over a conventional weight loss approach: a fast response, high-sensitivity detection, a small sample requirement, multiple sample measurements, and easy disassembly for cleaning and reuse purposes.

This paper describes the method of a novel impedance-based platform for the detection of the evaporation rate. The model compound hyaluronic acid was ...

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Video: A High Performance Impedance-based Platform for Evaporation Rate Detection

The Visual Colorimetric Detection of Multi-nucleotide Polymorphisms on a Pneumatic Droplet Manipulation Platform

A simple and visual method to detect multi-nucleotide polymorphism (MNP) was performed on a pneumatic droplet manipulation platform on an open surface. This approach to colorimetric DNA detection was based on the hybridization-mediated growth of gold nanoparticle probes (AuNP probes). The growth size and configuration of the AuNP are dominated by the number of DNA samples hybridized with the probes. Based on the specific size- and shape-dependent optical properties of the nanoparticles, the number of mismatches in a sample DNA fragment to the probes is able to be discriminated. The tests were conducted via droplets containing reagents and DNA samples respectively, and were transported and mixed on the pneumatic platform with the controlled pneumatic suction of the flexible PDMS-based superhydrophobic membrane. Droplets can be delivered simultaneously and precisely on an open-surface on the proposed pneumatic platform that is highly biocompatible with no side effect of DNA samples inside the droplets. Combining the two proposed methods, the multi-nucleotide polymorphism can be detected at sight on the pneumatic droplet manipulation platform; no additional instrument is required. The procedure from installing the droplets on the platform to the final result takes less than 5 min, much less than with existing methods. Moreover, this combined MNP detection approach requires a sample volume of only 10 &#181;l in each operation, which is remarkably less than that of a macro system.

This work presents a simple and visual method to detect multi-nucleotide polymorphisms on a pneumatic droplet manipulation platform. With the proposed method, the entire experiment, including droplet manipulation and detection of multi-nucleotide polymorphisms, can be performed near 23 &#176;C without the aid of advanced instruments.

A simple and visual method to detect multi-nucleotide polymorphism (MNP) was performed on a pneumatic droplet manipulation platform on an open surface. ...

Genetics

Exploring the Effects of Atmospheric Forcings on Evaporation: Experimental Integration of the Atmospheric Boundary Layer and Shallow Subsurface

Evaporation is directly influenced by the interactions between the atmosphere, land surface and soil subsurface. This work aims to experimentally study evaporation under various surface boundary conditions to improve our current understanding and characterization of this multiphase phenomenon as well as to validate numerical heat and mass transfer theories that couple Navier-Stokes flow in the atmosphere and Darcian flow in the porous media. Experimental data were collected using a unique soil tank apparatus interfaced with a small climate controlled wind tunnel. The experimental apparatus was instrumented with a suite of state of the art sensor technologies for the continuous and autonomous collection of soil moisture, soil thermal properties, soil and air temperature, relative humidity, and wind speed. This experimental apparatus can be used to generate data under well controlled boundary conditions, allowing for better control and gathering of accurate data at scales of interest not feasible in the field. Induced airflow at several distinct wind speeds over the soil surface resulted in unique behavior of heat and mass transfer during the different evaporative stages.

A protocol for the design and construction of a soil tank interfaced to a small climate controlled wind tunnel to study the effects of atmospheric forcings on evaporation is presented. Both the soil tank and wind tunnel are instrumented with sensor technologies for the continuous in situ measurement of environmental conditions.

Evaporation is directly influenced by the interactions between the atmosphere, land surface and soil subsurface. This work aims to experimentally study ...

Environment

A Simple, Low-cost, and Robust System to Measure the Volume of Hydrogen Evolved by Chemical Reactions with Aqueous Solutions

There is a growing research interest in the development of portable systems which can deliver hydrogen on-demand to proton exchange membrane (PEM) hydrogen fuel cells. Researchers seeking to develop such systems require a method of measuring the generated hydrogen. Herein, we describe a simple, low-cost, and robust method to measure the hydrogen generated from the reaction of solids with aqueous solutions. The reactions are conducted in a conventional one-necked round-bottomed flask placed in a temperature controlled water bath. The hydrogen generated from the reaction in the flask is channeled through tubing into a water-filled inverted measuring cylinder. The water displaced from the measuring cylinder by the incoming gas is diverted into a beaker on a balance. The balance is connected to a computer, and the change in the mass reading of the balance over time is recorded using data collection and spreadsheet software programs. The data can then be approximately corrected for water vapor using the method described herein, and parameters such as the total hydrogen yield, the hydrogen generation rate, and the induction period can also be deduced. The size of the measuring cylinder and the resolution of the balance can be changed to adapt the setup to different hydrogen volumes and flow rates.

The study of methods to generate on-demand hydrogen for fuel cells continues to grow in importance. However, systems to measure hydrogen evolution from the reaction of chemicals with water can be complicated and expensive. This article details a simple, low-cost, and robust method to measure the evolution of hydrogen gas.

There is a growing research interest in the development of portable systems which can deliver hydrogen on-demand to proton exchange membrane (PEM) ...

Chemistry

Electrochemical Impedance Spectroscopy as a Tool for Electrochemical Rate Constant Estimation

Electrochemical impedance spectroscopy (EIS) was used for advanced characterization of organic electroactive compounds along with cyclic voltammetry (CV). In the case of fast reversible electrochemical processes, current is predominantly affected by the rate of diffusion, which is the slowest and limiting stage. EIS is a powerful technique that allows separate analysis of stages of charge transfer that have different AC frequency response. The capability of the method was used to extract the value of charge transfer resistance, which characterizes the rate of charge exchange on the electrode-solution interface. The application of this technique is broad, from biochemistry up to organic electronics. In this work, we are presenting the method of analysis of organic compounds for optoelectronic applications.

Electrochemical impedance spectroscopy (EIS) of species that undergo reversible oxidation or reduction in solution was used for determination of rate constants of oxidation or reduction.

Electrochemical impedance spectroscopy (EIS) was used for advanced characterization of organic electroactive compounds along with cyclic voltammetry (CV). ...

Quantification and Size-profiling of Extracellular Vesicles Using Tunable Resistive Pulse Sensing

Extracellular vesicles (EVs), including &#8216;microvesicles&#8217; and &#8216;exosomes&#8217;, are highly abundant in bodily fluids. Recent years have witnessed a tremendous increase in interest in EVs. EVs have been shown to play important roles in various physiological and pathological processes, including coagulation, immune responses, and cancer. In addition, EVs have potential as therapeutic agents, for instance as drug delivery vehicles or as regenerative medicine. Because of their small size (50 to 1,000 nm) accurate quantification and size profiling of EVs is technically challenging.
This protocol describes how tunable resistive pulse sensing (tRPS) technology, using the qNano system, can be used to determine the concentration and size of EVs. The method, which relies on the detection of EVs upon their transfer through a nano sized pore, is relatively fast, suffices the use of small sample volumes and does not require the purification and concentration of EVs. Next to the regular operation protocol an alternative approach is described using samples spiked with polystyrene beads of known size and concentration. This real-time calibration technique can be used to overcome technical hurdles encountered when measuring EVs directly in biological fluids.

Extracellular vesicles play important roles in physiological and pathological processes, including coagulation, immune responses, and cancer or as potential therapeutic agents in drug delivery or regenerative medicine. This protocol presents methods for the quantification and size characterization of isolated and non-isolated extracellular vesicles in various fluids using tunable resistive pulse sensing.

Extracellular vesicles (EVs), including &#8216;microvesicles&#8217; and &#8216;exosomes&#8217;, are highly abundant in bodily fluids. Recent years have ...

Bioengineering

Protocol for Soil Hydraulic Property Measurement via Evaporation

Measurements of Soil Water Potential and Conductivity based on a Simple Evaporation Experiment using a Hydraulic Property Analyzer

The measurement of soil hydraulic properties is critical in understanding the physical components of soil health as well as integrated knowledge of soil systems under various management practices. Collecting reliable data is imperative for informing decisions that affect agriculture and the environment. The simple evaporation experiment described here uses instrumentation in a laboratory setting to analyze soil samples collected in the field. The soil water tension of the sample is measured by the instrument, and tension data is modeled by software to return soil hydraulic properties. This method can be utilized to measure soil water retention and hydraulic conductivity and give insight into differences in treatments or environmental dynamics over time. Initial establishment requires a user, but data acquisition is automated with the instrument. Soil hydraulic properties are not easily measured with traditional experiments, and this protocol offers a simple and optimal alternative. Interpretation of results and options for extending the data range are discussed.

Author Spotlight: Advancing Agricultural Land Ecosystem Research with a Hydraulic Property Analyzer to Assess Soil Health

This article features a simple evaporation experiment using a hydraulic property instrument for a soil sample. Through efficient means, measurements can be taken over a series of days to generate high-quality data.

The measurement of soil hydraulic properties is critical in understanding the physical components of soil health as well as integrated knowledge of soil ...

Studying Surfactant Effects on Hydrate Crystallization at Oil-Water Interfaces Using a Low-Cost Integrated Modular Peltier Device

We introduce an approach to study the formation and growth of hydrates under the influence of nonionic surfactants. The experimental system includes a temperature regulator, visualization techniques, and inner pressure measurements. The temperature control system contains a low-cost, programmable temperature regulator made with solid-state Peltier components. Along with the temperature control system, we incorporated visualization techniques and internal pressure measurements to study hydrate formation and inhibition in the presence of nonionic surfactants. We studied the hydrate-inhibiting ability of nonionic surfactants (sorbitane monolaurate, sorbitane monooleate, PEG-PPG-PEG, and polyoxyethylenesorbitan tristearate) at low (i.e., 0.1 CMC), medium (i.e., CMC), and high (i.e., 10 CMC) concentrations. Two types of crystals were formed: planar and conical. Planar crystals were formed in plain water and low surfactant concentrations. Conical crystals were formed in high surfactant concentrations. The results of the study show that conical crystals are the most effective in terms of hydrate inhibition. Because conical crystals cannot grow past a certain size, the hydrate growth rate as a conical crystal is slower than the hydrate growth rate as planar crystal. Hence, surfactants that force hydrates to form conical crystals are the most efficient. The goal of the protocol is to provide a detailed description of an experimental system that is capable of investigating the cyclopentane hydrate crystallization process on the surface of a water droplet in the presence of surfactant molecules.

We present a protocol to study the formation of hydrates in the presence of nonionic surfactants on the interface of a water droplet submerged in cyclopentane. The protocol consists of building a low-cost, programmable, temperature regulator. The temperature control system is combined with visualization techniques and internal pressure measurements.

We introduce an approach to study the formation and growth of hydrates under the influence of nonionic surfactants. The experimental system includes a ...

Enhancing EV Isolation for Precise Biofluid Analysis

Chemical Affinity-Based Isolation of Extracellular Vesicles from Biofluids for Proteomics and Phosphoproteomics Analysis

Extracellular vesicles (EVs) from biofluids have recently gained significant attention in the field of liquid biopsy. Released by almost every type of cell, they provide a real-time snapshot of host cells and contain a wealth of molecular information, including proteins, in particular those with post-translational modifications (PTMs) such as phosphorylation, as the main player of cellular functions and disease onset and progression. However, the isolation of EVs from biofluids remains challenging due to low yields and impurities from current EV isolation methods, making the downstream analysis of EV cargo, such as EV phosphoproteins, difficult. Here, we describe a rapid and effective EV isolation method based on functionalized magnetic beads for EV isolation from biofluids such as human urine and downstream proteomics and phosphoproteomics analysis following EV isolation. The protocol enabled a high recovery yield of urinary EVs and sensitive profiles of EV proteome and phosphoproteome. Furthermore, the versatility of this protocol and relevant technical considerations are also addressed here.

Author Spotlight: Advancing EVtrap for High-Throughput Proteomics in Disease Biomarker Discovery 

The present protocol provides detailed descriptions for the efficient isolation of urinary extracellular vesicles utilizing functionalized magnetic beads. Moreover, it encompasses subsequent analyses, including western blotting, proteomics, and phosphoproteomics.

Extracellular vesicles (EVs) from biofluids have recently gained significant attention in the field of liquid biopsy. Released by almost every type of ...

Biochemistry

A Detailed Protocol for Perspiration Monitoring Using a Novel, Small, Wireless Device

Perspiration monitoring can be utilized for the detection of certain diseases, such as thermoregulation and mental disorders, particularly when the patients are unaware of such disorders or are having difficulty expressing their symptoms. Until now, several devices for perspiration monitoring have been developed; however, such devices tend to have a relatively large exterior, considerable power consumption, and/or less sensitivity.
Recently, we developed a small, wireless device for perspiration monitoring. The device consists of a temperature/relative humidity (T/RH) sensor, battery-driven small data logger, and silica gel as a desiccant in a small cylindrical exterior. The T/RH sensor is placed between the detection windows (through which the water vapor from the skin enters) and the silica gel. The underlying principle of the perspiration monitoring device is based on Fick&#39;s law of diffusion, which means that water vapor flux from the skin to the silica gel (i.e.&#160;transepidermal water loss and perspiration) can be captured by change in humidity at the T/RH sensor. In addition, a baseline subtraction method was adopted to distinguish perspiration and transepidermal water loss.
As shown in the previous report, the developed device can monitor the perspiration at any sites of the body in an easy, wireless manner. However, detailed methods of how to use the device have not been disclosed yet. In this article, therefore, we would like to show the point-by-point tutorials of how to use the device for perspiration monitoring, by showing the sympathetic activity test with the sympathetic skin response monitoring as an example.

Recently, we developed a small wireless device for perspiration monitoring. In this article, we present detailed protocols on how to use the device for perspiration monitoring with an example of the sympathetic activity test.

Perspiration monitoring can be utilized for the detection of certain diseases, such as thermoregulation and mental disorders, particularly when the ...

Medicine

Measurement of Leaf Hydraulic Conductance and Stomatal Conductance and Their Responses to Irradiance and Dehydration Using the Evaporative Flux Method (EFM)

Water is a key resource, and the plant water transport system sets limits on maximum growth and drought tolerance. When plants open their stomata to achieve a high stomatal conductance (gs) to capture CO2 for photosynthesis, water is lost by transpiration1,2. Water evaporating from the airspaces is replaced from cell walls, in turn drawing water from the xylem of leaf veins, in turn drawing from xylem in the stems and roots. As water is pulled through the system, it experiences hydraulic resistance, creating tension throughout the system and a low leaf water potential (&Psi;leaf). The leaf itself is a critical bottleneck in the whole plant system, accounting for on average 30% of the plant hydraulic resistance3. Leaf hydraulic conductance (Kleaf = 1/ leaf hydraulic resistance) is the ratio of the water flow rate to the water potential gradient across the leaf, and summarizes the behavior of a complex system: water moves through the petiole and through several orders of veins, exits into the bundle sheath and passes through or around mesophyll cells before evaporating into the airspace and being transpired from the stomata. Kleaf is of strong interest as an important physiological trait to compare species, quantifying the effectiveness of the leaf structure and physiology for water transport, and a key variable to investigate for its relationship to variation in structure (e.g., in leaf venation architecture) and its impacts on photosynthetic gas exchange. Further, Kleaf responds strongly to the internal and external leaf environment3. Kleaf can increase dramatically with irradiance apparently due to changes in the expression and activation of aquaporins, the proteins involved in water transport through membranes4, and Kleaf declines strongly during drought, due to cavitation and/or collapse of xylem conduits, and/or loss of permeability in the extra-xylem tissues due to mesophyll and bundle sheath cell shrinkage or aquaporin deactivation5-10. Because Kleaf can constrain gs and photosynthetic rate across species in well watered conditions and during drought, and thus limit whole-plant performance they may possibly determine species distributions especially as droughts increase in frequency and severity11-14.
We present a simple method for simultaneous determination of Kleaf and gs on excised leaves. A transpiring leaf is connected by its petiole to tubing running to a water source on a balance. The loss of water from the balance is recorded to calculate the flow rate through the leaf. When steady state transpiration (E, mmol &bull; m-2 &bull; s-1) is reached, gs is determined by dividing by vapor pressure deficit, and Kleaf by dividing by the water potential driving force determined using a pressure chamber (Kleaf= E /- &Delta;&psi;leaf, MPa)15.
This method can be used to assess Kleaf responses to different irradiances and the vulnerability of Kleaf to dehydration14,16,17.

Measurement of Leaf Hydraulic Conductance and Stomatal Conductance and Their Responses to Irradiance and Dehydration Using the Evaporative Flux Method EFM

We describe a relatively rapid (30 min) and realistic method for simultaneously measurement of leaf hydraulic conductance (Kleaf) and stomatal conductance (gs) for transpiring excised leaves. The method can be modified to measure the light and dehydration responses of Kleaf and gs.

Water is a key resource, and the plant water transport system sets limits on maximum growth and drought tolerance. When plants open their stomata to ...

A High Performance Impedance-based Platform for Evaporation Rate Detection

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