• Clinical and Experimental Reproductive Medicine
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• 제27권1호
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• pp.47-57
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• 2000

Objective: The effects of cryopreservation with or without coculture on the in vitro development of blastomere-biopsied 8-cell mouse embryos were investigated. This experimental study was originally designed for the setup of a preclinical mouse model for the preimplantation genetic diagnosis (PGD) in human. Methods: Eight-cell embryos were obtained after in vitro fertilization (IVF) from F1 hybrid mice (C57BL(표현불가)/CBA(표현불가)). Using micromanipulation, one to four blastomeres were aspirated through a hole made in the zona pellucida by zona drilling (ZD) with acid Tyrode's solution (ATS). A slow-freezing and rapid-thawing protocol with 1.5M dimethyl sulfoxide (DMSO) and 0.1M sucrose as cryoprotectant was used for the cryopreservation of blastomere- biopsied 8-cell mouse embryos. After thawing, embryos were cultured for 110 hours in Ham's F-10 supplemented with 0.4% bovine serum albumin (BSA). In the coculture group, embryos were cultured for 110 hours on the monolayer of Vero cells in the same medium. The blastocyst formation was recorded, and the embryos developed beyond blastocyst stage were stained with 10% Giemsa to count the total number of nuclei in each embryo. Results: The survival rate of embryos after cryopreservation was significantly lower in the blastomere-biopsied (7/8, 6/8, 5/8, and 4/8 embryos) groups than in the non-biopsied, zona intact (ZI) group. Without the coculture, the blastocyst formation rate of embryos after cryopreservation was not significantly different among ZI, the zona drilling only (ZD), and the balstomere-biopsied groups, but it was significantly lower than in the non-cryopreserved control group. The mean number of cells in embryos beyond blastocyst stage was significantly higher in the control group ($50.2{\pm}14.0$) than in 6/8 ($26.5{\pm}6.2$), 5/8 ($25.0{\pm}5.5$), and 4/8 ($17.8{\pm}7.8$) groups. With the coculture using Vero cells, the blastocyst formation rate of embryos after cryopreservation was significantly lower in 5/8 and 4/8 groups, compared with the control, 7/8, and 6/8 groups. The mean number of cells in embryos beyond blastocyst stage was also significantly lower in 4/8 group ($25.9{\pm}10.2$), compared with the control ($50.2{\pm}14.0$), 7/8 ($56.0{\pm}22.2$), and 6/8 ($55.3{\pm}25.5$) groups. Conclusion: After cryopreservation, blastomere-biopsied mouse embryos have a significantly impaired developmental competence in vitro, but this detrimental effect might be prevented by the coculture with Vero cells in 8-cell mouse embryos biopsied one or two blastomeres. Biopsy of mouse embryos after ZD with ATS is a safe and highly efficient preclinical model for PGD of human embryos.

• 정신신체의학
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• 제28권2호
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• pp.185-193
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• 2020

연구목적 본 연구의 목적은 한국판 섭식장애 진단척도 DSM-5 (Korean version Eating Disorder Diagnostic Scale, K-EDDS)를 웹 기반 진단 시스템으로 개발하고 타당도를 검증하는 데 있다. 방 법 본 연구는 섭식장애 환자(38명)와 대학생(81명)을 포함하여 총 119명이 참여하였다. 모든 참가자는 지필 섭식장애 스크리닝 검사인 Sick, Control, One, Fat, Food (SCOFF) 후 별도의 사이트에서 웹 기반 K-EDDS, 섭식장애검사(Eating Disorder Examination-Questionnaire, EDE-Q), 임상손상평가(Clinical Impairment Assessment Questionnaire, CIA)를 작성했다. SCOFF 점수가 2점 이상인 사람을 대상으로 EDE (Eating Disorder Examination Interview) 면담을 진행하였다. 검사 후 2주 이내에 웹 기반 K-EDDS, EDE-Q, CIA를 재실시하였다. 결 과 탐색적 요인분석 결과, 신체불만족, 폭식행동, 폭식빈도, 보상행동의 4가지 요인이 추출되어 총 분산의 82.4%를 설명하였다. 웹 기반 K-EDDS의 4개 하위요인은 EDE-Q의 4개 하위요인과 각각 유의미한 상관관계가 있었으며, 우수한 내적 일치도를 보였다(Cronbach's alpha=0.93). 웹 기반 K-EDDS와 EDE의 진단 일치도는 96.83%, 웹 기반 K-EDDS의 검사-재검사 진단 일치도는 92.86%로 우수하였다. 웹 기반 K-EDDS와 CIA에서 환자군과 정상군 간 차이가 유의하게 나타나, 본 척도의 판별 타당도를 검증하였다. 결 론 본 연구를 통해 웹 기반 K-EDDS는 임상 및 연구 현장에서 DSM-5를 기반으로 한 섭식장애 진단에 유용하게 사용할 수 있는 타당한 도구임을 확인하였다.

• 대한바이러스학회지
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• 제26권2호
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• pp.215-225
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• 1996

Aseptic meningits, an acute inflammation of the meninges, is a common illness during childhood. Virus is the most important cause of aseptic meningitis. Especially enterovirus causes approximately above 85% of all cases of aseptic meningitis. In 1993, there was a big epidemic of aseptic meningitis by ECHO 9 and ECHO 30 viruses. And ECHO 3 virus was isolated as a causative agent of aseptic meningitis in 1994. This study was aimed to detect the causative agent of aseptic meningitis in 1995 and to analyze the 5'-noncoding region which was used to detect virus. Virus was isolated from 87 stools and cerebrospinal fluid specimens of the patients by cultured RD and HEp-2 cell. Neutralizing antibody tests using enterovirus serum pool were performed on the specimens with cytopathic effect. 3 of ECHO 7 viruses and 5 of Coxsackie B3 viruses were isolated from stool specimens and 1 of ECHO 7 and Coxsackie B3 mixed type was confirmed from cerebrospinal fluid specimens. RNA was isolated from the culture supernatants of infected cells and general primers were selected in highly conserved part of the 5'-noncoding region of the enteroviral genome for RT-PCR. PCR product from this virus showed a 152bp band on gel electrophoresis. Sequence of obtained DNA was compared with prototype sequences by accessing to the Genebank database. 5'-noncoding region of isolated Coxsackie B3 virus, which has point mutations in nucleotide sequence positions 493, 497, 502, 523, was closely related to that of polio virus type 1, Mahoney strain. In case of isolated ECHO 7 virus, nucleotide has been changed from cytosine to thymine at position 581 and from thymine to cytosine at position 583. We concluded the causative agents of the outbreak of aseptic meningitis during June to July in 1995 were both ECHO 7 and Coxsackie B3 virus, and the primer used in this study could allow a rapid diagnosis of enteroviruses by PCR.

• Parasites, Hosts and Diseases
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• 제50권1호
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• pp.15-21
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• 2012

In Iran, $Plasmodium$ $vivax$ is responsible for more than 80% of the infected cases of malaria per year. Control interventions for vivax malaria in humans rely mainly on developed diagnostic methods. Recombinant $P.$ $vivax$ apical membrane antigen-1 (rPvAMA-1) has been reported to achieve designing rapid, sensitive, and specific molecular diagnosis. This study aimed to perform isolation and expression of a rPvAMA-1, derived from Iranian patients residing in an endemic area. Then, the diagnostic efficiency of the characterized Iranian PvAMA-1 was assessed using an indirect ELISA method. For this purpose, a partial region of AMA-1 gene was amplified, cloned, and expressed in pET32a plasmid. The recombinant $His-tagged$ protein was purified and used to coat the ELISA plate. Antibody detection was assessed by indirect ELISA using rPvAMA-1. The validity of the ELISA method for detection of anti-$P.$ $vivax$ antibodies in the field was compared to light microscopy on 84 confirmed $P.$ $vivax$ patients and compared to 84 non-$P.$ $vivax$ infected individuals. The ELISA cut-off value was calculated as the mean+2SD of OD values of the people living in malaria endemic areas from a south part of Iran. We found a cut-off point of OD=0.311 that showed the best correlation between the sera confirmed with $P.$ $vivax$ infection and healthy control sera. A sensitivity of 81.0% and specificity of 84.5% were found at this cut off titer. A good degree of statistical agreement was found between ELISA using rPvAMA-1 and light microscopy (0.827) by Kappa analysis.

• Journal of Microbiology and Biotechnology
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• 제26권1호
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• pp.180-189
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• 2016

The widespread occurrence of drug-resistant Mycobacterium tuberculosis places importance on the detection of TB (tuberculosis) drug susceptibility. Conventional drug susceptibility testing (DST) is a lengthy process. We developed a rapid enzymatic color-reaction-based biochip assay. The process included asymmetric multiplex PCR/templex PCR, biochip hybridization, and an enzymatic color reaction, with specific software for data operating. Templex PCR (tem-PCR) was applied to avoid interference between different primers in conventional multiplex-PCR. We applied this assay to 276 clinical specimens (including 27 sputum, 4 alveolar lavage fluid, 2 pleural effusion, and 243 culture isolate specimens; 40 of the 276 were non-tuberculosis mycobacteria specimens and 236 were M. tuberculosis specimens). The testing process took 4.5 h. A sensitivity of 50 copies per PCR was achieved, while the sensitivity was 500 copies per PCR when tem-PCR was used. Allele sequences could be detected in mixed samples at a proportion of 10%. Detection results showed a concordance rate of 97.46% (230/236) in rifampicin resistance detection (sensitivity 95.40%, specificity 98.66%) and 96.19% (227/236) in isoniazid (sensitivity 93.59%, specificity 97.47%) detection with those of DST assay. Concordance rates of testing results for sputum, alveolar lavage fluid, and pleural effusion specimens were 100%. The assay provides a potential choice for TB diagnosis and treatment.

• 생명과학회지
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• 제25권8호
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• pp.903-909
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• 2015

LAMP (Loop-mediated Isothermal Amplification)법은 PCR를 기반으로 등온에서 autocycling 가닥 변위 DNA 합성에 의존하며, Bst polymerase를 사용하여 진단하는 방법이다. 이것은 대상 DNA의 여섯 개의 배열을 인식하는 4개의 특정 primer의 도움을 받아 단시간 안에 병원체를 식별하는 높은 특이성을 지니고 있다. 본 연구에서는 LAMP로 수생에서 위험한 병원체인 Vibrio alginolyticus의 특별한 LAMP primer를 제작하였으며, 신속한 진단을 위해 MgSO₄, dNTP, Betaine, Bst polymerase의 최적 반응 조건의 특이성 및 기존의 PCR보다 10배 정도의 민감하다는 것을 확인하였다. 또한, 디자인 되어진 LAMP primer가 다른 Vibrio 종들 중 오직 V. alginolyticus에서만 반응한 것을 확인 할 수 있었다. 본 논문에서는 병원체 세균인 V. alginolyticus의 빠르고 민감한 효과적인 진단으로 양식 질병들을 조기에 발견할 수 있도록 개발하였다.

• 동국한의학연구소논문집
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• 제7권2호
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• pp.141-148
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• 1999

FoLT-PCR 기술을 인체체질의학적 응용을 위하여 당뇨병연구에 사용하였다. 당뇨병은 제1형 및 제2형으로 나뉘는데 제1형은 인슈린비의존성(NIDDM)으로 당뇨병환자의 약60%이상을 차지하며, 제2형은 인슈린의존성(IDDM)으로 당뇨병환자의 30%미만을 차지한다. 이들은 대부분 후천적으로 환경중에서의 인슈린관련 유전자의 돌연변이에 의해 발병하는 것으로 알려져 있다. 이에, 본 연구에서는 당뇨병의 병인을 유전적변이와 체질의학적 관계에서 고찰하기 위하여 수행되었다. NIDDM환자와 IDDM환자를 대상으로 인슈린유전자를 증폭하여 제한효소절단 양상과 염기배열분석을 하였다. 비암호영역중 4개 위치 +216, +1045, +1367, 및 +1380에서 다형성을 보였으며 새로운 ${\alpha}4$, ${\alpha}5$, ${\alpha}6$ 및 ${\beta}2$이 ${\alpha}1$과 ${\beta}1$가 이형(heterozygous)에서만 검출되며 ${\alpha}1$은 우성이며 신규형들과 ${\beta}1$은 열성이었다. 이러한 당뇨병병인은 유전학적으로 체질의학과 깊은 관계를 가지는 것을 시사하였다.

• 한국미생물·생명공학회지
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• 제43권3호
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• pp.296-299
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• 2015

Cowpea mild mottle virus (CPMMV)는 국내 미보고 바이러스로, 넓은 숙주범위를 가지며, 국내 관리급 검역바이러스로 지정되어 있다. 본 연구에서는 검역현장에서 신속하게 CPMMV를 진단할 수 있는 방법을 개발하였다. 이번 연구는 사용자를 위한 검사방법 개발을 위하여, 기존의 검역현장에서 활용하는 PCR 조성과 조건을 활용하였다. 본 연구에서는 CPMMV를 특이적으로 진단할 수 있는 2개의 RT-PCR 프라이머를 개발하였으며 각각 579와 638 bp를 증폭할 수 있다. 최종적으로 증폭되는 nested PCR 산물의 크기는 (579 → 298 bp)과 (638 → 252 bp)로 CPMMV의 특이 유전자를 진단할 수 있다. 또한, 제한효소 Xho I이 반응할 수 있는 염기 서열을 삽입하여 양성대조구 플라스미드를 개발하였다. 이것은 실험실 오염으로부터 거짓양성을 검증할 수 있게 하였다. 본 연구는 CPMMV와 관련된 수입 작물로부터 검역진단에 기여할 것이라고 사료된다.

• 한국산학기술학회논문지
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• 제17권6호
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• pp.469-481
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• 2016

최근 우리나라는 가족공동체의 해체 및 고령화 사회 진전, 주거문화의 변화로 1인가구가 급증하고 있어, 주거선택에 있어 일반 가구와는 다른 양상을 보이고 있다. 이에 1인가구 급증에 따른 주택시장의 변화를 진단하고 올바른 정책방향을 제시하는 방안 모색이 필요하다. 이 연구는 성인 1인 단독가구를 대상으로 이들의 주거점유형태에 영향을 미치는 요인이 무엇인지를 밝히고자 함에 그 목적이 있다. 분석은 한국복지패널 제 10차 조사자료를 이용하여 다항 로짓 모형을 통한 횡단면 분석을 실시하였으며, 그 분석 결과는 다음과 같다. 첫째, 20-30대 1인가구는 40대 이상보다 가구 및 주거특성이 더욱 열악하게 나타났다. 둘째, 20-30대 1인가구는 주택 점유형태를 선택할 때 영향을 미치는 가구의 특성은 40대 이상과 같은 특성에 대해서도 다른 방향의 결과를 보였다. 셋째, 20-30대 1인가구의 특성은 공공의 지원을 통해 개선의 여지를 보였다. 이 연구를 통해 정부의 주거 지원이 보다 더 다양한 방식으로 모색되어야 할 필요가 있음을 시사 해 주었다.

• Tuberculosis and Respiratory Diseases
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• 제79권4호
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• pp.282-288
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• 2016

Background: Tuberculosis (TB) is a major health problem, and accurate and rapid diagnosis of multidrug-resistant (MDR) and extended drug-resistant (XDR) TB is important for appropriate treatment. In this study, performances of solid and liquid culture methods were compared with respect to MDR- and XDR-TB isolate recovery and drug susceptibility testing. Methods: Sputum specimens from 304 patients were stained with Ziehl-Neelsen method. Mycobacterium tuberculosis (Mtb) isolates were tested for recovery on $L{\ddot{o}wenstein$-Jensen (LJ) medium and the BacT Alert 3D system. For drug susceptibility testing of Mtb, isolates were evaluated on M-KIT plates and the BacT Alert 3D system. Results: The recovery rates were 94.9% (206/217) and 98.2% (213/217) for LJ medium and the BacT Alert 3D system, respectively (kappa coefficient, 0.884). The rate of drug resistance was 13.4% for at least one or more drugs, 6.0% for MDR-TB and 2.3% for XDR-TB. M-KIT plate and BacT 3D Alert 3D system were comparable in drug susceptibility testing for isoniazid (97.7%; kappa coefficient, 0.905) and rifampin (98.6%; kappa coefficient, 0.907). Antibiotic resistance was observed using M-KIT plates for 24 of the total 29 Mtb isolates (82.8%). Conclusion: The liquid culture system showed greater reduction in the culture period, as compared with LJ medium; however, drug susceptibility testing using M-KIT plates was advantageous for simultaneous testing against multiple drug targets.