• Journal of IKEEE
• /
• v.23 no.1
• /
• pp.295-301
• /
• 2019

A heater in a portable real-time polymerase chain reaction(PCR) system is one of the important factors for controlling the PCR thermocycle precisely. Since heaters are integrated on a small-sized PCR chip for rapid heating and fabricated by semiconductor processes, the cost of producing PCR chips is high. Here, we propose to use chip resistors as an inexpensive and accurate temperature control method. The temperature distribution was simulated using one or two chip resistors on a real-time PCR chip and the PCR chip with uniform temperature distribution was fabricated. The temperature rise and fall rates were $18^{\circ}C/s$ and $3^{\circ}C/s$, respectively.

• Korean Journal of Agricultural Science
• /
• v.38 no.4
• /
• pp.761-767
• /
• 2011

The objective of this study was to predict total bacteria count of pork meats by using the portable electronic nose systems developed throughout two stages of the prototypes. Total bacteria counts were measured for pork meats stored at $4^{\circ}C$ for 21days and compared with the signals of the electronic nose systems. PLS(Partial least square), PCR (Principal component regression), MLR (Multiple linear regression) models were developed for the prediction of total bacteria count of pork meats. The coefficient of determination ($R_p{^2}$) and root mean square error of prediction (RMSEP) for the models were 0.789 and 0.784 log CFU/g with the 1st system for the pork loin, 0.796 and 0.597 log CFU/g with the 2nd system for the pork belly, and 0.661 and 0.576 log CFU/g with the 2nd system for the pork loin respectively. The results show that the developed electronic system has potential to predict total bacteria count of pork meats.

• Parasites, Hosts and Diseases
• /
• v.59 no.1
• /
• pp.77-82
• /
• 2021

As malaria remains a major health problem worldwide, various diagnostic tests have been developed, including microscopy-based and rapid diagnostic tests. LabChip real-time PCR (LRP) is a small and portable device used to diagnose malaria using lab-on-a-chip technology. This study aimed to evaluate the diagnostic performance of LRP for detecting malaria parasites. Two hundred thirteen patients and 150 healthy individuals were enrolled from May 2009 to October 2015. A diagnostic detectability of LRP for malaria parasites was compared to that of conventional RT-PCR. Sensitivity of LRP for Plasmodium vivax, P. falciparum, P. malariae, and P. ovale was 95.5%, 96.0%, 100%, and 100%, respectively. Specificity of LRP for P. vivax, P. falciparum, P. malariae, and P. ovale was 100%, 99.3%, 100%, and 100%, respectively. Cohen's Kappa coefficients between LRP and CFX96 for detecting P. vivax, P. falciparum, P. malariae, and P. ovale were 0.96, 0.98, 1.00, and 1.00, respectively. Significant difference was not observed between the results of LRP and conventional RT-PCR and microscopic examination. A time required to amplify DNAs using LRP and conventional RT-PCR was 27 min and 86 min, respectively. LRP amplified DNAs 2 times more fast than conventional RT-PCR due to the faster heat transfer. Therefore, LRP could be employed as a useful tool for detecting malaria parasites in clinical laboratories.

• Research in Plant Disease
• /
• v.18 no.4
• /
• pp.298-303
• /
• 2012

Tomato yellow leaf curl virus (TYLCV), transmitted exclusively by the whitefly (Bemisia tabaci) in a circulative manner is one of the most important virus in tomato. Since the first report of TYLCV incidence in Korea in 2008, the virus has rapidly spread nationwide. TYLCV currently causes serious economic losses in tomato production in Korea. Early detection of TYLCV is one of the most important methods to allow rouging of infected tomato plants to minimize the spread of TYLCV disease. We have developed an ultra-rapid and sensitive real-time polymerase chain reaction (PCR) using a new designed real-time PCR system, GenSpectorTM TMC-1000 that is a small and portable real-time PCR machine requiring only a $5{\mu}l$ reaction volume on microchips. The new system provides ultra-high speed reaction (30 cycles in less than 15 minutes) and melting curve analysis for amplified TYLCV products. These results suggest that the short reaction time and ultra sensitivity of the GenSpector$^{TM}$-based real-time PCR technique is suitable for monitoring epidemics and pre-pandemic TYLCV disease. This is the first report for plant virus detection using an ultra-rapid real-time PCR system.

• Analytical Science and Technology
• /
• v.29 no.2
• /
• pp.79-84
• /
• 2016

A small and inexpensive thermal cycler (PCR machine), known as the MiniPCR^TM Mini8 Thermal Cycler (Amplyus, Cambridge, MA, USA), was developed. In this study, the performance of this PCR machine was compared with the GeneAmp^® PCR system 9700 (Applied Biosystems) using four autosomal short tandem repeat (STR) kits, a Y-chromosome STR kit, and a mitochondrial DNA HV1/HV2 sequence analysis. The sensitivity and stochastic effects of the STR multiplex kits and the quality of the DNA sequence analysis were similar between the two PCR machines. The MiniPCR^TM Mini8 Thermal Cycler could be used for analyses at forensic DNA laboratories and crime scenes. The cost of the PCR is so economical that school laboratories and individuals could use the machines.

• Proceedings of the Korean Vacuum Society Conference
• /
• 2013.08a
• /
• pp.273-273
• /
• 2013

Recently, Point-of-care (POC) testing microdevices enable to do the patient monitoring, drug screening, pathogen detection in the outside of hospital. Immunochromatographic strip (ICS) is one of the diagnostic technologies which are widely applied to POC detection. Relatively low cost, simplicity to use, easy interpretations of the diagnostic results and high stability under any circumstances are representative advantages of POC diagnosis. It would provide colorimetric results more conveniently, if the genetic analysis microsystem incorporates the ICS as a detector part. In this work, we develop a reverse transcriptase-polymerase chain reaction (RT-PCR) microfluidic device integrated with a ROSGENE strip for colorimetric influenza H1N1 virus detection. The integrated RT-PCR- ROSGENE device is consist of four functional units which are a pneumatic micropump for sample loading, 2 ${\mu}L$ volume RT-PCR chamber for target gene amplification, a resistance temperature detector (RTD) electrode for temperature control, and a ROSGENE strip for target gene detection. The device was fabricated by combining four layers: First wafer is for RTD microfabrication, the second wafer is for PCR chamber at the bottom and micropump channel on the top, the third is the monolithic PDMS, and the fourth is the manifold for micropump operation. The RT-PCR was performed with subtype specific forward and reverse primers which were labeled with Texas-red, serving as a fluorescent hapten. A biotin-dUTP was used to insert biotin moieties in the PCR amplicons, during the RT-PCR. The RT-PCR amplicons were loaded in the sample application area, and they were conjugated with Au NP-labeled hapten-antibody. The test band embedded with streptavidins captures the biotin labeled amplicons and we can see violet colorimetric signals if the target gene was amplified with the control line. The off-chip RT-PCR amplicons of the influenza H1N1 virus were analyzed with a ROSGENE strip in comparison with an agarose gel electrophoresis. The intensities of test line was proportional to the template quantity and the detection sensitivity of the strip was better than that of the agarose gel. The test band of the ROSGENE strip could be observed with only 10 copies of a RNA template by the naked eyes. For the on-chip RT-PCR-ROSGENE experiments, a RT-PCR cocktail was injected into the chamber from the inlet reservoir to the waste outlet by the micro-pump actuation. After filling without bubbles inside the chamber, a RT-PCR thermal cycling was executed for 2 hours with all the microvalves closed to isolate the PCR chamber. After thermal cycling, the RT-PCR product was delivered to the attached ROSGENE strip through the outlet reservoir. After dropping 40 ${\mu}L$ of an eluant buffer at the end of the strip, the violet test line was detected as a H1N1 virus indicator, while the negative experiment only revealed a control line and while the positive experiment a control and a test line was appeared.

• Journal of Microbiology and Biotechnology
• /
• v.28 no.11
• /
• pp.1928-1936
• /
• 2018

Recently, human infections caused by severe fever with thrombocytopenia syndrome virus (SFTSV), which can lead to fatality, have dramatically increased in East Asia. With the unavailability of vaccines or antiviral drugs to prevent and/or treat SFTSV infection, early rapid diagnosis is critical for prevention and control of the disease. Here, we report the development of a simple, rapid and sensitive portable detection method for SFTSV infection applying reverse transcription-loop mediated isothermal amplification (RT-LAMP) combined with one-pot colorimetric visualization and electro-free reaction platform. This method utilizes a pocket warmer to facilitate diagnosis in a resource-limited setting. Specific primers were designed to target the highly-conserved region of L gene of SFTSV. The detection limit of the RT-LAMP assay was approximately $10^0$ viral genome copies from three different SFTSV strains. This assay exhibited comparable sensitivity to qRT-PCR and 10-fold more sensitivity than conventional RT-PCR, with a rapid detection time of 30 to 60 minutes. The RT-LAMP assay using SFTSV clinical specimens has demonstrated a similar detection rate to qRT-PCR and a higher detection rate compared to conventional RT-PCR. Moreover, there was no observed cross-reactive amplification of other human infectious viruses including Japanese Encephalitis Virus (JEV), Dengue, Enterovirus, Zika, Influenza and Middle East Respiratory Syndrome Coronavirus (MERS-CoV). This highly sensitive, electro- and equipment-free rapid colorimetric visualization method is feasible for resource-limited SFTSV field diagnosis.

• Proceedings of the Korean Vacuum Society Conference
• /
• 2013.08a
• /
• pp.88-89
• /
• 2013

A variety of influenza A viruses from animal hosts are continuously prevalent throughout the world which cause human epidemics resulting millions of human infections and enormous industrial and economic damages. Thus, early diagnosis of such pathogen is of paramount importance for biomedical examination and public healthcare screening. To approach this issue, here we propose a fully integrated Rotary genetic analysis system, called Rotary Genetic Analyzer, for on-site detection of influenza A viruses with high speed. The Rotary Genetic Analyzer is made up of four parts including a disposable microchip, a servo motor for precise and high rate spinning of the chip, thermal blocks for temperature control, and a miniaturized optical fluorescence detector as shown Fig. 1. A thermal block made from duralumin is integrated with a film heater at the bottom and a resistance temperature detector (RTD) in the middle. For the efficient performance of RT-PCR, three thermal blocks are placed on the Rotary stage and the temperature of each block is corresponded to the thermal cycling, namely $95^{\circ}C$ (denature), $58^{\circ}C$ (annealing), and $72^{\circ}C$ (extension). Rotary RT-PCR was performed to amplify the target gene which was monitored by an optical fluorescent detector above the extension block. A disposable microdevice (10 cm diameter) consists of a solid-phase extraction based sample pretreatment unit, bead chamber, and 4 ${\mu}L$ of the PCR chamber as shown Fig. 2. The microchip is fabricated using a patterned polycarbonate (PC) sheet with 1 mm thickness and a PC film with 130 ${\mu}m$ thickness, which layers are thermally bonded at $138^{\circ}C$ using acetone vapour. Silicatreated microglass beads with 150~212 ${\mu}L$ diameter are introduced into the sample pretreatment chambers and held in place by weir structure for construction of solid-phase extraction system. Fig. 3 shows strobed images of sequential loading of three samples. Three samples were loaded into the reservoir simultaneously (Fig. 3A), then the influenza A H3N2 viral RNA sample was loaded at 5000 RPM for 10 sec (Fig. 3B). Washing buffer was followed at 5000 RPM for 5 min (Fig. 3C), and angular frequency was decreased to 100 RPM for siphon priming of PCR cocktail to the channel as shown in Figure 3D. Finally the PCR cocktail was loaded to the bead chamber at 2000 RPM for 10 sec, and then RPM was increased up to 5000 RPM for 1 min to obtain the as much as PCR cocktail containing the RNA template (Fig. 3E). In this system, the wastes from RNA samples and washing buffer were transported to the waste chamber, which is fully filled to the chamber with precise optimization. Then, the PCR cocktail was able to transport to the PCR chamber. Fig. 3F shows the final image of the sample pretreatment. PCR cocktail containing RNA template is successfully isolated from waste. To detect the influenza A H3N2 virus, the purified RNA with PCR cocktail in the PCR chamber was amplified by using performed the RNA capture on the proposed microdevice. The fluorescence images were described in Figure 4A at the 0, 40 cycles. The fluorescence signal (40 cycle) was drastically increased confirming the influenza A H3N2 virus. The real-time profiles were successfully obtained using the optical fluorescence detector as shown in Figure 4B. The Rotary PCR and off-chip PCR were compared with same amount of influenza A H3N2 virus. The Ct value of Rotary PCR was smaller than the off-chip PCR without contamination. The whole process of the sample pretreatment and RT-PCR could be accomplished in 30 min on the fully integrated Rotary Genetic Analyzer system. We have demonstrated a fully integrated and portable Rotary Genetic Analyzer for detection of the gene expression of influenza A virus, which has 'Sample-in-answer-out' capability including sample pretreatment, rotary amplification, and optical detection. Target gene amplification was real-time monitored using the integrated Rotary Genetic Analyzer system.

• Journal of Microbiology and Biotechnology
• /
• v.33 no.5
• /
• pp.559-573
• /
• 2023

Shiga toxin (Stxs)-producing enterohaemorrhagic Escherichia coli (EHEC) and Shigella dysenteriae serotype 1 are major causative agents of severe bloody diarrhea (known as hemorrhagic colitis) and hemolytic uremic syndrome (HUS) associated with extraintestinal complications such as acute renal failure and neurologic impairment in infected patients under 9 years of age. Extreme nephrotoxicity of Stxs in HUS patients is associated with severe outcomes, highlighting the need to develop technologies to detect low levels of the toxin in environmental or food samples. Currently, the conventional polymerase chain reaction (PCR) or immunoassay is the most broadly used assay to detect the toxin. However, these assays are laborious, time-consuming, and costly. More recently, numerous studies have described novel, highly sensitive, and portable methods for detecting Stxs from EHEC. To contextualize newly emerging Stxs detection methods, we briefly explain the basic principles of these methods, including lateral flow assays, optical detection, and electrical detection. We subsequently describe existing and newly emerging rapid detection technologies to identify and measure Stxs.

• Food Engineering Progress
• /
• v.13 no.4
• /
• pp.297-301
• /
• 2009

This study was performed to develop a prediction model of sugar content for strawberry. Near-infrared (NIR) spectroscopy has been prevailed for on-line and portable applications for non-invasive quality assessment of intact fruit. This work presents effects of illumination method and coating of reflection surface of light source on prediction result of sugar content. Effect of preprocessing methods was also examined. A low-cost commercially available VIS/NIR spectrometer was used for estimation of total soluble solids content (Brix). To predict sugar contents of strawberry, the best results were obtained with the spectrum data measured under intensive illuminations at three locations induced from the light source with fiber optic bundles. Gold coating of reflection surface of light source lamp gave favorable effect to prediction result. The best results in validation of PLSR model were $r_{SEP}$ = 0.891 and SEP = 0.443 Brix under OSC preprocessing and those of PCR were $r_{SEP}$ = 0.845, SEP $r_{SEP}$= 0.520 Brix, under no preprocessing.