• Title/Summary/Keyword: In-Vitro Diagnostic

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Application of Engineered Zinc Finger Proteins Immobilized on Paramagnetic Beads for Multiplexed Detection of Pathogenic DNA

  • Shim, Jiyoung;Williams, Langley;Kim, Dohyun;Ko, Kisung;Kim, Moon-Soo
    • Journal of Microbiology and Biotechnology
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    • v.31 no.9
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    • pp.1323-1329
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    • 2021
  • Micro-scale magnetic beads are widely used for isolation of proteins, DNA, and cells, leading to the development of in vitro diagnostics. Efficient isolation of target biomolecules is one of the keys to developing a simple and rapid point-of-care diagnostic. A zinc finger protein (ZFP) is a double-stranded (ds) DNA-binding domain, providing a useful scaffold for direct reading of the sequence information. Here, we utilized two engineered ZFPs (Stx2-268 and SEB-435) to detect the Shiga toxin (stx2) gene and the staphylococcal enterotoxin B (seb) gene present in foodborne pathogens, Escherichia coli O157 and Staphylococcus aureus, respectively. Engineered ZFPs are immobilized on a paramagnetic bead as a detection platform to efficiently isolate the target dsDNA-ZFP bound complex. The small paramagnetic beads provide a high surface area to volume ratio, allowing more ZFPs to be immobilized on the beads, which leads to increased target DNA detection. The fluorescence signal was measured upon ZFP binding to fluorophore-labeled target dsDNA. In this study, our system provided a detection limit of ≤ 60 fmol and demonstrated high specificity with multiplexing capability, suggesting a potential for development into a simple and reliable diagnostic for detecting multiple pathogens without target amplification.

Peripheral Blood Lymphocytes as In Vitro Model to Evaluate Genomic Instability Caused by Low Dose Radiation

  • Tewari, Shikha;Khan, Kainat;Husain, Nuzhat;Rastogi, Madhup;Mishra, Surendra P;Srivastav, Anoop K
    • Asian Pacific Journal of Cancer Prevention
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    • v.17 no.4
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    • pp.1773-1777
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    • 2016
  • Diagnostic and therapeutic radiation fields are planned so as to reduce side-effects while maximising the dose to site but effects on healthy tissues are inevitable. Radiation causes strand breaks in DNA of exposed cells which can lead to chromosomal aberrations and cause malfunction and cell death. Several researchers have highlighted the damaging effects of high dose radiation but still there is a lacuna in identifying damage due to low dose radiation used for diagnostic purposes. Blood is an easy resource to study genotoxicity and to estimate the effects of radiation. The micronucleus assay and chromosomal aberration can indicate genetic damage and our present aim was to establish these with lymphocytes in an in vitro model to predict the immediate effects low dose radiation. Blood was collected from healthy individuals and divided into 6 groups with increasing radiation dose i.e., 0Gy, 0.10Gy, 0.25Gy, 0.50Gy, 1Gy and 2Gy. The samples were irradiated in duplicates using a LINAC in the radiation oncology department. Standard protocols were applied for chromosomal aberration and micronucleus assays. Metaphases were stained in Giemsa and 200 were scored per sample for the detection of dicentric or acentric forms. For micronuclei detection, 200 metaphases. Giemsa stained binucleate cells per sample were analysed for any abnormality. The micronuclei (MN) frequency was increased in cells exposed to the entire range of doses (0.1-2Gy) delivered. Controls showed minimal MN formation ($2.0%{\pm}0.05$) with triple MN ($5.6%{\pm}2.0$) frequency at the lowest dose. MN formation increased exponentially with the radiation dose thereafter with a maximum at 2Gy. Significantly elevated numbers of dicentric chromosomes were also observed, even at doses of 0.1-0.5Gy, compared to controls, and acentric chromosomes were apparent at 2Gy. In conclusion we can state that lymphocytes can be effectively used to study direct effect of low dose radiation.

Effect of object position in the field of view and application of a metal artifact reduction algorithm on the detection of vertical root fractures on cone-beam computed tomography scans: An in vitro study

  • Nikbin, Ava;Kajan, Zahra Dalili;Taramsari, Mehran;Khosravifard, Negar
    • Imaging Science in Dentistry
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    • v.48 no.4
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    • pp.245-254
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    • 2018
  • Purpose: To assess the effects of object position in the field of view (FOV) and application of a metal artifact reduction (MAR) algorithm on the diagnostic accuracy of cone-beam computed tomography (CBCT) for the detection of vertical root fractures(VRFs). Materials and Methods: Sixty human single-canal premolars received root canal treatment. VRFs were induced in 30 endodontically treated teeth. The teeth were then divided into 4 groups, with 2 groups receiving metal posts and the remaining 2 only having an empty post space. The roots from different groups were mounted in a phantom made of cow rib bone, and CBCT scans were obtained for the 4 different groups. Three observers evaluated the images independently. Results: The highest frequency of correct diagnoses of VRFs was obtained with the object positioned centrally in the FOV, using the MAR algorithm. Peripheral positioning of the object without the MAR algorithm yielded the highest sensitivity for the first observer (66.7%). For the second and third observers, a central position improved sensitivity, with or without the MAR algorithm. In the presence of metal posts, central positioning of the object in the FOV significantly increased the diagnostic sensitivity and accuracy compared to peripheral positioning. Conclusion: Diagnostic accuracy was higher with central positioning than with peripheral positioning, irrespective of whether the MAR algorithm was applied. However, the effect of the MAR algorithm was more significant with central positioning than with peripheral positioning of the object in the FOV. The clinical experience and expertise of the observers may serve as a confounder in this respect.

Detection of proximal caries using quantitative light-induced fluorescence-digital and laser fluorescence: a comparative study

  • Yoon, Hyung-In;Yoo, Min-Jeong;Park, Eun-Jin
    • The Journal of Advanced Prosthodontics
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    • v.9 no.6
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    • pp.432-438
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    • 2017
  • PURPOSE. The purpose of this study was to evaluate the in vitro validity of quantitative light-induced fluorescence-digital (QLF-D) and laser fluorescence (DIAGNOdent) for assessing proximal caries in extracted premolars, using digital radiography as reference method. MATERIALS AND METHODS. A total of 102 extracted premolars with similar lengths and shapes were used. A single operator conducted all the examinations using three different detection methods (bitewing radiography, QLF-D, and DIAGNOdent). The bitewing x-ray scale, QLF-D fluorescence loss (${\Delta}F$), and DIAGNOdent peak readings were compared and statistically analyzed. RESULTS. Each method showed an excellent reliability. The correlation coefficient between bitewing radiography and QLF-D, DIAGNOdent were -0.644 and 0.448, respectively, while the value between QLF-D and DIAGNOdent was -0.382. The kappa statistics for bitewing radiography and QLF-D had a higher diagnosis consensus than those for bitewing radiography and DIAGNOdent. The QLF-D was moderately to highly accurate (AUC = 0.753 - 0.908), while DIAGNOdent was moderately to less accurate (AUC = 0.622 - 0.784). All detection methods showed statistically significant correlation and high correlation between the bitewing radiography and QLF-D. CONCLUSION. QLF-D was found to be a valid and reliable alternative diagnostic method to digital bitewing radiography for in vitro detection of proximal caries.

High Prevalence of Fluoroquinolone- and Methicillin-Resistant Staphylococcus pseudintermedius Isolates from Canine Pyoderma and Otitis Externa in Veterinary Teaching Hospital

  • Yoo, Jong-Hyun;Yoon, Jang-W.;Lee, So-Young;Park, Hee-Myung
    • Journal of Microbiology and Biotechnology
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    • v.20 no.4
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    • pp.798-802
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    • 2010
  • Recently, a total of 74 Staphylococcus pseudintermedius isolates were collected from clinical cases of canine pyoderma and otitis externa in Korea. In this study, we examined in vitro fluoroquinolone resistance among those isolates using a standard disc diffusion technique. The results demonstrated that, except for one isolate, approximately 18.9% to 27.0% of the isolates possessed bacterial resistance to both veterinary- and human-licensed fluoroquinolones including moxifloxacin (18.9% resistance), levofloxacin (20.3% resistance), ofloxacin (24.3% resistance), ciprofloxacin (25.7% resistance), and enrofloxacin (27.0% resistance). Most surprisingly, 14 out of 74 (18.9%) isolates were resistant to all the five fluoroquinolones evaluated. Moreover, a PCR detection of the methicillin resistance gene (mecA) among the 74 isolates revealed that 13 out of 25 (52.0%) mecApositive isolates, but only 7 out of 49 (14.3%) mecA-negative isolates, were resistant to one or more fluoroquinones. Taken together, our results imply that bacterial resistance to both veterinary- and human-use fluoroquinolones becomes prevalent among the S. pseudintermedius isolates from canine pyoderma and otitis externa in Korea, as well as that the high prevalence of the mecA-positive S. pseudintermedius isolates carrying multiple fluoroquinolones resistance could be a potential public health problem.

In Vitro Imaging of MRI and Ultrasound for Gastric Carcinoma (위암 조직의 자기공명영상과 초음파 소견에 대한 비교 연구)

  • Kil, Sung-Won;Jee, Keum-Nahn
    • Investigative Magnetic Resonance Imaging
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    • v.12 no.2
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    • pp.178-187
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    • 2008
  • Purpose : To evaluate and compare the diagnostic accuracy of MRI and ultrasound(US) for estimation of invasion depth of gastric carcinoma by correlation with histopathologic findings in vitro and to find out the best MR pulse sequence for detection and accurate delineation of tumor. Materials and Methods : Resected specimen of total or subtotal gastrectomy from 53 patients with gastric carcinoma were done of imaging studies of MRI and US. And US was examined by using high frequency linear transducer for tumor invasion depth by a radiologist. In each case, both imaging findings of MRI and US were evaluated independently for tumor detection and invasion depth by consensus of two radiologists and were compared the diagnostic accuracy between two imaging modalities according to the histopathologic findings. MR imaging with five MR pulse sequences, spin echo T1 and in- and out-of phase gradient echo T1 weighted images, FSE and SSFSE T2 weighted images, were performed. Five MR pulse sequences were evaluated and compared on the point of detection and accurate distinction of tumor from surrounding normal tissue. Results : In EGC, diagnostic accuracy of US(77%) was superior than that of MRI(59%) but no statistically significant difference was noted between two imaging modalities(p=0.096). In AGC, both imaging modalities of MRI and US showed relatively high diagnostic accuracy as 97% and 84% respectively. Diagnostic accuracy of MRI was statistically better than that of US at the significant level(p<0.001). The best MR pulse sequence among five in each specimen was FSE T2WI(75.5%, 40/53) in both EGC and AGC. In AGC, FSE T2WI showed excellent imaging quality by showing very high ratio (93.5%, 29/31) of accurate delineation of tumor. Conclusion : MRI and US show relatively high diagnostic accuracy in the evaluation of tumor invasion depth of resected specimen in AGC. The most excellent pulse sequence of MRI for the evaluation of tumor invasion depth is FSE T2WI on the point of detection and accurate delineation of tumor in both EGC and AGC.

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The effects of diagnostic hysteroscopy on the reproductive outcomes of infertile women without intrauterine pathologies: a systematic review and meta-analysis

  • Yang, Soo Yeon;Chon, Seung-Joo;Lee, Seon Heui
    • Women's Health Nursing
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    • v.26 no.4
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    • pp.300-317
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    • 2020
  • Purpose: Hysteroscopy can be used both to diagnose and to treat intrauterine pathologies. It is well known that hysteroscopy helps to improve reproductive outcomes by treating intrauterine pathologies. However, it is uncertain whether hysteroscopy is helpful in the absence of intrauterine pathologies. This study aimed to confirm whether hysteroscopy improves the reproductive outcomes of infertile women without intrauterine pathologies. Methods: We conducted a systematic review of 11 studies retrieved from Ovid-MEDLINE, Ovid-Embase, and the Cochrane Library. Two independent investigators extracted the data and used risk-of-bias tools (RoB 2.0 and ROBINS-I) to assess their quality. Results: Diagnostic hysteroscopy prior to in vitro fertilization (IVF)/intracytoplasmic sperm injection (ICSI) was associated with a higher clinical pregnancy rate (CPR) and live birth rate (LBR) than non-hysteroscopy in patients with recurrent implantation failure (RIF) (odds ratio, 1.79 and 1.46; 95% confidence interval, 1.40-2.30 and 1.08-1.97 for CPR and LBR, respectively) while hysteroscopy prior to first IVF was ineffective. The overall meta-analysis of LBR showed statistically significant findings for RIF, but a subgroup analysis showed effects only in prospective cohorts (odds ratio, 1.40 and 1.47; 95% confidence interval, 0.62-3.16 and 1.04-2.07 for randomized controlled trials and prospective cohorts, respectively). Therefore, the LBR should be interpreted carefully and further research is needed. Conclusion: Although further research is warranted, hysteroscopy may be considered as a diagnostic and treatment option for infertile women who have experienced RIF regardless of intrauterine pathologies. This finding enables nurses to educate and support infertile women with RIF prior to IVF/ICSI.

Comparison of fluorescence loss measurements among various generations of QLF devices (세대별 QLF 기기의 평활면 비와동형 법랑질 우식 병소 탐지에 관한 진단정확도 비교)

  • Park, Seok-Woo;Lee, Hyung-Suk;Kim, Sang-Kyeom;Lee, Eun-Song;de Jong, Elbert de Josselin;Kim, Baek-Il
    • The Journal of the Korean dental association
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    • v.56 no.1
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    • pp.8-16
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    • 2018
  • Purpose: The aim of in vitro study was to compare the diagnostic accuracy to detect non-cavitated enamel caries on smooth surface by using four kinds of the QLF devices. Materials and Methods: A total of 52 human permanent premolars and molars were used. Fluorescence images were captured by the QLF devices (Inspektor Pro, QLF-D, Qraycam, and Qraypen). Fluorescence loss of the QLF was calculated. The severity of lesions was categorized into the following 3 scores using polarized light microscopy: normal (S), enamel demineralization to outer half of enamel (D1), and inner half of the enamel up to the dentin-enamel junction (D2). The Kruskal-Wallis test was used to compare the fluorescence loss among the QLF devices. Spearman rank correlation coefficient between histological scores and fluorescence loss of the devices was calculated. The sensitivity, specificity, and area under the receiver operating curve (AUROC) were calculated to compare their diagnostic accuracies. Results: The correlation coefficients between histological scores and the fluorescence loss of the devices showed 0.77 to 0.81 (P < 0.001). All histological scores, the fluorescence loss among the devices showed no statistical difference. Among the devices, sensitivity, specificity, and AUC values of the fluorescence loss showed 0.84 to 0.94, 0.76 to 0.90, and 0.90 to 0.92, respectively. Conclusions: All QLF devices had no difference with excellent diagnostic accuracies to detect non-cavitated enamel caries on smooth surface.

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Clinical In Vivo Bio Assay of Glucose in Human Skin by a Tattoo Film Carbon Nano Tube Sensor

  • Ly, Suw Young;Lee, Chang Hyun
    • Journal of the Korean Applied Science and Technology
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    • v.34 no.3
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    • pp.595-601
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    • 2017
  • In vivo assay of glucose detection was described using a skin tattoo film electrode (STF), and the probe was made from carbon nano tube paste modification film paper. Here in the square-wave stripping anodic working range obtained of $20-100mgL^{-1}$ within an accumulation time of 0 seconds only in sea water electrolyte solutions of pH 7.0. The relative standard deviations of 50 mg glucose that were observed of 0.14 % (n=12), respectively, using optimum stripping accumulation of 30 sec, the low detection limit (S/N) was pegged at 15.8 mg/L. The developed results can be applied to the detect of in vivo skin sensing in real time. Which confirms the results are usable for in vitro or vivo diagnostic clinical analysis.

Allergy tests in clinical practice (알레르기 검사의 실제)

  • Lee, Jaechun
    • Journal of Medicine and Life Science
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    • v.15 no.1
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    • pp.1-5
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    • 2018
  • Allergy is conditions when a hypersensitivity reaction happens with a certain element, called as an allergen, which is commonly not reactive to ordinary individuals. Allergic diseases involve various organs or systems in the body. The purpose of allergy tests is to make a diagnosis of allergic diseases and to identify the affecting allergens. In vivo tests, more relevant in clinical situation, include skin test, patch test and provocation test. In in vitro tests, there are specific IgE test, histamine releasing assay, and lymphocyte activation test, safer and more objective than in vivo tests. In the view point of clinical practice, skin test, provocation test, total IgE test and specific IgE test were reviewed in depth.