• Parasites, Hosts and Diseases
• /
• v.52 no.3
• /
• pp.263-271
• /
• 2014

PCR detection of intestinal protozoa is often restrained by a poor DNA recovery or by inhibitors present in feces. The need for an extraction protocol that can overcome these obstacles is therefore clear. $QIAamp^{(R)}$ DNA Stool Mini Kit (Qiagen) was evaluated for its ability to recover DNA from oocysts/cysts directly from feces. Twenty-five Giardia-positive, 15 Cryptosporidium-positive, 15 Entamoeba histolytica-positive, and 45 protozoa-free samples were processed as control by microscopy and immunoassay tests. DNA extracts were amplified using 3 sets of published primers. Following the manufacturer's protocol, the kit showed sensitivity and specificity of 100% towards Giardia and Entamoeba. However, for Cryptosporidium, the sensitivity and specificity were 60% (9/15) and 100%, respectively. A series of optimization experiments involving various steps of the kit's protocol were conducted using Cryptosporidium-positive samples. The best DNA recoveries were gained by raising the lysis temperature to the boiling point for 10 min and the incubation time of the InhibitEX tablet to 5 min. Also, using a pre-cooled ethanol for nucleic acid precipitation and small elution volume ($50-100{\mu}l$) were valuable. The sensitivity of the amended protocol to Cryptosporidium was raised to 100%. Cryptosporidium DNA was successfully amplified by either the first or the second primer set. When applied on parasite-free feces spiked with variable oocysts/cysts counts, ${\approx}2$ oocysts/cysts were theoretically enough for detection by PCR. To conclude, the Qiagen kit with the amended protocol was proved to be suitable for protozoan DNA extraction directly from feces and support PCR diagnosis.

• Journal of Chest Surgery
• /
• v.33 no.8
• /
• pp.669-675
• /
• 2000

Background: Tuberculous pleurisy is the leading cause of pleural effusion in Korea. And differential diagnosis of tuberculous pleurisy with other cause is clinically very important. Traditional diagnostic methods such as routine analysis of pleural fluid, staining for acid-fast bacilli or pleural biopsy have major inherent limitaion. This study was designed to evaluate the significance of pleural fluid polymerase chain reaction(PCR) and adenosine deaminase (ADA) activity in early diagnosis of tuberculous pleurisy. Material and Method: Between March 1996 and July 1997, 198 patients with pleural effusion reviewed retrospectively. The study group included 112 cases with tuberculous effusion and 86 cases with non-tuberculous effusions, whose diagnoses were confirmed by pleural biopsy, microbiological methods, or cytology. We compared the results of PCR and pleural fluid levels of ADA between tuberculous and non-tuberculous effusions. Result: Mean age was 47.54$\pm$19.52 years(range 2 to 85 years). The positive rate of PCR was significantly higher in tuberculous group than non-tuberculous group(p<0.05). The sensitivty, specificity, positive predictive value(PPV), and negative predictive value(NPV) for PCR were 31.7, 90.9, 83.0, and 48.8%, respectively. Mean ADA activity was significantly higher in tuberculous group than non-tuberculous group(83.2 U/L vs 49.8 U/L)(p<0.05). With diagnostic thresholds of 40 U/L, the sensitivity, specificity, PPV, and NPV of ADA for tuberculosis were 75.9, 70.9, 77.3, and 69.3% respectively. At a level of 70 U/L, the sensitivity, specificity, PPV, and NPV of ADA for tuberculosis were 70.1, 75.9, 82.9, and 60.3% respectively. Conclusion: PCR is very highly specific, but less sensitive methods in diagnosis of tuberculous pleurisy. But ADA level of pleural fluid has acceptable sensitivity and specificity in diagnosis of tuberculous pleurisy. ADA activity is more useful test in the evaluation of pleural effusions.

• Proceedings of the KIEE Conference
• /
• 2005.07c
• /
• pp.2410-2412
• /
• 2005

In this study, two types of PS substrate were fabricated for sensing of chemical and biological substances. For sensing of the humidity and chemical analyzes such as $CH_3OH$ or $C_2H_5OH$, PS layers are prepared by photoelectrochemical etching of silicon wafer in aqueous hydrofluoric acid solution. To evaluate their sensitivity, we measured the resistance variation of the PS diaphragm. As the amplitude of applied voltage increases from 2 to 6Vpp at constant frequency of 5kHz, the resistance variation for humidity sensor rises from 376.3 to $784.8{\Omega}$/%RH. And the sensitivities for $CH_3OH$ and $C_2H_5OH$ were 0.068 uA/% and 0.212 uA/%, respectively. For biological sensing application, amperometric urea sensors were fabricated based on porous silicon(PS), and planar silicon(PLS) electrode substrates by the electrochemical methods. Pt thin film was sputtered on these substrates which were previously formed by electrochemical anodization. Poly (3-methylthiophene) (P3MT) were used for electron transfer matrix between urease(Urs) and the electrode phase, and Urs also was by electrochemically immobilized. Effective working area of these electrodes was determined for the first time by using $Fe(CN)_6^{3-}/Fe(CN)_6^{4-}$ redox couple in which nearly reversible cyclic voltammograms were obtained. The $i_p$ vs $v^{1/2}$ plots show that effective working electrode area of the PS-based Pt thin film electrode was 1.6 times larger than the PLS-based one and we can readily expect the enlarged surface area of PS electrode would result in increased sensitivity by ca. 1.6 times. Actually, amperometric sensitivity of the Urs/P3MT/Pt/PS electrode was ca 0.91uA/$mM{\cdot}cm^2$, and that of the Urs/P3MT/Pt/PLS electrode was ca. 0.91uA/$mM{\cdot}cm^2$ in a linear range of 1mmol/L to 100mmol/L urea concentrations

• The Korean Journal of Physiology and Pharmacology
• /
• v.25 no.5
• /
• pp.479-488
• /
• 2021

This study aimed to develop docetaxel (DTX) loaded poly(lactic-co-glycolic acid) (PLGA) nanoparticles (DTX-NPs) and to evaluate the different pharmacological sensitivity of NPs to MCF-7 and MDA-MB-231 breast cancer cells. NPs containing DTX or coumarin-6 were prepared by the nanoprecipitation method using PLGA as a polymer and d-α-tocopherol polyethylene glycol 1000 succinate (TPGS) as a surfactant. The physicochemical properties of NPs were characterized. In vitro anticancer effect and cellular uptake were evaluated in breast cancer cells. The particle size and zeta potential of the DTX-NPs were 160.5 ± 3.0 nm and -26.7 ± 0.46 mV, respectively. The encapsulation efficiency and drug loading were 81.3 ± 1.85% and 10.6 ± 0.24%, respectively. The in vitro release of DTX from the DTX-NPs was sustained at pH 7.4 containing 0.5% Tween 80. The viability of MDA-MB-231 and MCF-7 cells with DTX-NPs was 37.5 ± 0.5% and 30.3 ± 1.13%, respectively. The IC₅₀ values of DTX-NPs were 3.92- and 6.75-fold lower than that of DTX for MDA-MB-231 cells and MCF-7 cells, respectively. The cellular uptake of coumarin-6-loaded PLGA-NPs in MCF-7 cells was significantly higher than that in MDA-MB-231 cells. The pharmacological sensitivity in breast cancer cells was higher on MCF-7 cells than on MDA-MB-231 cells. In conclusion, we successfully developed DTX-NPs that showed a great potential for the controlled release of DTX. DTX-NPs are an effective formulation for improving anticancer effect in breast cancer cells.

• Biomedical Science Letters
• /
• v.17 no.3
• /
• pp.191-196
• /
• 2011

Noroviruses (noroV) are the major cause of nonbacterial gastroenteritis in humans worldwide. Since noroV cannot yet be cultured in vitro and their diagnosis by electron microscopy requires at least $10^6$ viral particles/g of stool a variety of molecular detection techniques represent an important step towards the detection of noroV. In the present study, we have applied real-time nucleic acid sequence-based amplification (real-time NASBA) for simultaneous detection of NoroV genogroup I (GI) and genogroup II (GII) using standard viral RNA. For real-time NASBA assay which can detected noroV GI and GII, a selective region of the genes encoding the capsid protein was used to design primers and genotype-specific molecular beacon probes. The specificity of the real-time NASBA using newly designed primers and probes were confirmed using standard viral RNA of noroV GI and GII. To determine the sensitivity of this assay, serial 10-fold dilutions of standard viral RNA of noroV GI and GII were used for reverse transcription polymerase chain reaction (RT-PCR) and real-time NASBA. The results showed that while agarose gel electrophoresis could detect RT-PCR products with 10 pg of standard viral RNA, the real-time NASBA assay could detect 100 fg of standard viral RNA. These results suggested that the real-time NASBA assay has much higher sensitivity than conventional RT-PCR assay. This assay was expected that might detect the viral RNA in the specimens which could have been false negative by RT-PCR. There were needed to perform real-time NASBA with clinical specimens for evaluating accurate sensitivity and specificity of this assay.

• Biomedical Science Letters
• /
• v.26 no.3
• /
• pp.170-178
• /
• 2020

Mycobacterium tuberculosis (MTB) continues to be one of the main causative agents of tuberculosis (TB); moreover, the incidence of nontuberculous mycobacteria (NTM) infections has been rising gradually in both immunocompromised and immunocompetent patients. Precise and rapid detection and identification of MTB and NTM in respiratory specimens are thus important for MTB infection control. Molecular diagnostic methods based on the nucleic acid amplification test (NAAT) are known to be rapid, sensitive, and specific compared to the conventional acid-fast bacilli (AFB) smear and mycobacterial culture methods. In the present study, the clinical performances of three commercial molecular diagnostic assays, namely TB/NTM PCR (Biocore), MolecuTech Real MTB-ID^® (YD Diagnostics), and REBA Myco-ID^® (YD Diagnostics), were evaluated with a total of 92 respiratory specimens (22 AFB smear positives and 67 AFB smear negatives). The sensitivity and specificity of TB/NTM PCR were 100% and 75.81%, respectively. The corresponding values of MolecuTech Real MTB-ID^® and REBA Myco-ID^® were 56.52% and 90.32%, and 56.52% and 82.26%, respectively. TB/NTM PCR showed the highest sensitivity; however, the concordant rate was 10% compared with sequence analysis. Although MolecuTech Real MTB-ID^® showed lower sensitivity, its specificity was the highest among the three methods. REBA Myco-ID^® allowed accurate classification of NTM species; therefore, it was the most specific diagnostic method. Of the three PCR-based methods, MolecuTech Real MTB-ID^® showed the best performance. This method is expected to enable rapid and accurate identification of MTB and NTM.

• Asian Pacific Journal of Cancer Prevention
• /
• v.16 no.14
• /
• pp.5889-5897
• /
• 2015

Background: The performance of combined testing visual inspection with acetic acid (VIA) and cervical cytology tests might differ from one setting to another. The average estimate of the testing accuracy across studies is informative, but no meta-analysis has been carried out to assess this combined method. Objective: The objective of this study was to estimate the average sensitivity and specificity of the combined VIA and cervical cytology tests for the detection of cervical precancerous lesions. Materials and Methods: We conducted a systematic review and a meta-analysis, according to the Cochrane Handbook for Systematic Review of Diagnostic Test Accuracy. We considered two cases. In the either-positive result case, a positive result implies positivity in at least one of the tests. A negative result implies negativity in both tests. In the both-positive case, a positive result implies having both tests positive. Eligible studies were identified using Pubmed, Embase, Website of Science, CINHAL and COCRANE databases. True positive, false positive, false negative and true negative values were extracted. Estimates of sensitivity and specificity, positive and negative likelihood (LR) and diagnostic odds ratios (DOR) were pooled using a hierarchical random effect model. Hierarchical summary receiver operating characteristics (HSROC) were generated and heterogeneity was verified through covariates potentially influencing the diagnostic odds ratio. Findings: Nine studies fulfilled inclusion criteria and were included in the analysis. Pooled estimates of the sensitivities of the combined tests in either-positive and both-positive cases were 0.87 (95% CI: 0.83-0.90) and 0.38 (95% CI: 0.29-0.48), respectively. Corresponding specificities were 0.79 (95% CI: 0.63-0.89) and 0.98 (95% CI: 0.96-0.99) respectively. The DORs of the combined tests in either-positive or both-positive result cases were 27.7 (95% CI: 12.5-61.5) and 52 (95% CI: 22.1-122.2), respectively. When including only articles without partial verification bias and also a high-grade cervical intraepithelial neoplasia as a threshold of the disease, DOR of combined test in both-positive result cases remained the highest. However, DORs decreased to 12.1 (95% CI: 6.05-24.1) and 13.8 (95% CI: 7.92-23.9) in studies without partial verification bias for the combined tests in the either-positive and both-positive result cases, respectively. The screener, the place of study and the size of the population significantly influenced the DOR of combined tests in the both-positive result case in restriction analyses that considered only articles with CIN2+ as disease threshold. Conclusions: The combined test in the either-positive result case has a high sensitivity, but a low specificity. These results suggest that the combined test should be considered in developing countries as a primary screening test if facilities exist to confirm, through colposcopy and biopsy, a positive result.

• Investigative Magnetic Resonance Imaging
• /
• v.25 no.3
• /
• pp.172-182
• /
• 2021

Purpose: We evaluated the diagnostic performance of LI-RADS version 2018 using gadoxetic acid enhanced MRI for recurrent but untreated HCC in patients with prior history of HCC. Materials and Methods: We enrolled 50 consecutive patients who 1) prior history of treatment of HCC, 2) underwent liver surgery for radiological/clinical diagnosis of new HCC between 2013 to 2018, 3) had gadoxetic acid enhanced MRI within one month before surgery, and 4) did not have more than five HCCs or infiltrative tumors only. Two radiologists reviewed MRI and determined the presence of LR3, LR4 and LR5 observations except previously treated tumors based on LI-RADS version 2018 in consensus. We sub-classified LR4 into LR4m (LR4 with major features only) and LR4u (LR4 upgraded from LR3 by ancillary features). LR4u were further sub-classified into LR4ua (with arterial phase hyperenhancement) and LR4un (without arterial phase hyperenhancement). Results: PPV for LR5, LR4 and LR3 observations for recurrent HCC were 100%, 61.5% and 25.0%, respectively. 100% (3/3) of LR4m were HCC. However, PPV of LR4u was 56.5%. PPV of LR4ua and LR4un were 73.3% and 25.0%, respectively. Sensitivity of LR5 and LR5+LR4 observations as a diagnostic threshold were 32.1% and 89.3%, respectively. Sensitivity for LR5+LR4m+LR4ua observations for diagnosis of HCC were 83.7% and significantly superior to that of LR5 without significant deterioration of specificity (75.0%). Conclusion: In patients with prior history of HCC, LR4 observations by major features or with APHE may be regarded as recurrent HCCs given high sensitivity and comparable specificity/PPV to LR5 observations.

• Applied Chemistry for Engineering
• /
• v.24 no.5
• /
• pp.471-475
• /
• 2013

This study is about photosensitive microspheres prepared by coating alginate microspheres with gelatin-cinnamic acid conjugate. Firstly, alginate microspheres was prepared in water-in-oil (W/O) emulsion and then they were coated with gelatin- cinnamic acid conjugate. Herein, gelatin-cinnamic acid conjugate is obtained by the amidation between an amine group of gelatin and a carboxy group of cinnamic acid. Cinnamic acid is widely used as a photo-responsive material easy to dimerize and dedimeriz under UV irradiation at ${\lambda}$ = 254 nm and ${\lambda}$ = 365 nm, respectively. As shown in SEM-EDS, alginate was successfully coated with gelatin-ciannmic acid. By determining the absorbance of coated microspheres at 270nm, the amount of cinnamic acid per microspheres was 0.13/1. The SEM photos showed the size of coated microspheres is around $10{\mu}m$. And the degrees of dimerization and dedimerization were calculated to be 49% and 23% respectively. Then the release of FITC-dextran from the coated micrspheres was studied and release the degree was 42%. As a result, the coated microspheres have potential to be used as a photo-responsive drug carrier to delivery drugs.

• Journal of Electrochemical Science and Technology
• /
• v.10 no.4
• /
• pp.361-372
• /
• 2019

The silver nanoparticles/polyaniline/reduced graphene oxide nanocomposite modified glassy carbon electrode (Ag/PANI/RGO/GCE) was prepared by the electrochemical method. The Ag/PANI/RGO nanocomposite was characterized by transmission electron microscopy (TEM), field emission scanning electron microscopy (FE-SEM), Raman spectroscopy, X-ray diffraction (XRD), and electrochemical impedance spectroscopy (ESI). Two electrochemical techniques namely differential pulse voltammetry (DPV) and cyclic voltammetry (CV) were used to the electrochemical behaviors investigation of ascorbic acid (AA), dopamine (DA), and uric acid (UA). The Ag/PANI/RGO/GCE exhibited remarkable electrocatalytic activity towards the oxidation reaction of AA, DA, and UA in Britton-Robinson (BR) solution (pH=4.0). Under the optimal conditions, the determinations of AA, DA, and UA were accomplished using DPV. AA-DA and DA-UA peak potential separations were 130 and 180 mV, respectively. For simultaneous detection, the linear response ranges were in the two concentration ranges of 0.05-0.8 mM and 2.0-16.0 mM with detection limit 0.412 μM (S/N = 3) for AA, 0.7-90.0 μM and 90.0-1000.0 μM with detection limit 0.023 μM (S/N = 3) for DA, and 0.8-70.0 μM and 70.0-1000.0 μM with detection limit 0.050 μM (S/N = 3) for UA. This modified electrode showed good sensitivity, selectivity, and stability with applied to determine AA, DA, and UA in human urine and drug.