• 대한임상검사과학회지
• /
• 제46권2호
• /
• pp.47-53
• /
• 2014

The purpose of this study was to evaluate the clinical utility of TB/NTM PCR by comparing the results of TB PCR to detect Mycobacterium tuberculous (MTB) and nontuberculous mycobacteria (NTM) from paraffin-embedded tissue specimens. A total of 60 cases were tested using TB PCR and TB/NTM PCR. The MTB and NTM rate of TB/NTM PCR was 84.2% (16/19), 10.5% (2/19) in TB positive of TB PCR. The NTM rate of TB/NTM PCR was 29.3% (12/41) in TB negative of TB PCR. Fourteen different species of NTM were identified, the common isolate was M. gordonae (21.4%), M. avium (14.3%), M. ulcerans (7.1%), M. interjectum (7.1%), M. gilvum (7.1%), M. fortuitum (7.1%), M. mucogenicum (7.1%). The rare species identified were M. farcinogenes (7.1%), M. tokaiense (7.1%). Therefore, TB/NTM PCR is useful to differentiate MTB and NTM from paraffin-embedded tissue specimens and it is more effective in detecting NTM with TB PCR.

• Advances in pediatric surgery
• /
• 제11권1호
• /
• pp.9-18
• /
• 2005

Abnormal distribution of enteric nerves such as adrenergic, cholinergic and non-adrenergic non-cholinergic nerves (NANC) may cause the failure of relaxation at the involved bowel segment in Hirschsprung's disease (HD). Nitric oxide (NO) is a major inhibitory NANC neurotransmitter in the gastrointestinal tract. NO is synthesized by activation of nNOS (neuronal nitric oxide synthase) in the intramural ganglion cells and regulates bowel movement. To assess the distribution of nNOS in HD, immunohistochemical staining to nNOS was utilized on paraffin embedded specimens. Ten control colon specimens were tested for feasibility of staining. Immunohistochemisrty was done on ganglionic colon as well as aganglionic segment of 15 patients with HD. nNOS immunoreactivity was observed in the neuronal cells, small cells and nerve fibers in the muscle layer and submucosal neuronal cells of control specimens. This finding was also observed in the ganglionic segments of HD. But, there was no nNOS immunoreactivity in aganglionic segments of HD. In conclusion nNOS immunohistochemical staining of paraffin embedded specimen is feasible and reliable. And the results suggest that the relaxation failure of the aganglionic bowel in HD is related to the absence of nNOS containing cells and nerve fibers.

• 대한의생명과학회지
• /
• 제6권1호
• /
• pp.55-63
• /
• 2000

PCR을 이용한 임상가검물 중에서 보편화된 객담 이외에 미량의 각종 체액과 임상에서 많이 실시하지 않는 파라핀 포매 조직에서의 결핵균 검출을 실험하여 그 활용 가능성을 규명하고자 하였다. 임상가검물인 체액 65예는 항산성 염색과 배양검사, PCR을 실시하였고, 파라핀 포매 조직 50예는 항산성 염색과 병리조직학적 진단, PCR을 실시하여 다음과 같은 결론을 얻었다. 본 실험 검체 중 임상가검물인 체액에서 항산성 염색 음성인 검체 중 12.1%,배양검사에서 음성인 검체 중 3.7%에서 PCR 양성의 결과를 보였고, 파라핀 포매 조직에서는 항산성 염색 음성인 검체 중 20.0%에서 PCR양성의 결과를 얻어 PCR이 민감도와 특이도가 높음을 확인할 수 있었다.

• Asian Pacific Journal of Cancer Prevention
• /
• 제15권6호
• /
• pp.2733-2737
• /
• 2014

Objective: Molecular pathology tests are often carried for clinicopathological diagnosis and pathologists have established large collections of formalin-fixed, paraffin-embedded tissue (FFPE) banks. However, extraction of DNA from FFPE is a laborious and challenging for researchers in clinical laboratories. The aim of this study was to compare two widely used DNA extraction methods: using a QIAamp DNA FFPE kit from Qiagen and a Cobas Sample Preparation Kit from Roche, and evaluated the effect of the DNA quality on molecular diagnostics. Methods: DNA from FFPE non-small cell lung carcinoma tissues including biopsy and surgical specimens was extracted with both QIAamp DNA FFPE and Cobas Sample Preparation Kits and EGFR mutations of non-small cell lung carcinomas were detected by real-time quantitative PCR using the extracted DNA. Results and Conclusion: Our results showed that DNA extracted by QIAamp and Cobas methods were both suitable to detect downstream EGFR mutation in surgical specimens. Howover, Cobas method could yield more DNA from biopsy specimens, and gain much better EGFR mutation results.

• Journal of Genetic Medicine
• /
• 제2권2호
• /
• pp.53-57
• /
• 1998

Spinal muscular atrophy (SMA) type I is a common severe autosomal recessive inherited neuromuscular disorder that has been mapped to chromosome 5q11.2-13.3. The survival motor neuron (SMN) gene, a candidate gene, is known to be deleted in 96% of patients with SMA type I. Presently, PCR and single strand conformation polymorphism (PCR-SSCP) analyses have been made possible for application to both archival slides and paraffin-embedded tissues. Archival materials represent valuable DNA resources for genetic diagnosis. We applied these methods for the identification of SMN gene of SMA type I in archival specimens for the prenatal diagnosis. In this study, we performed the prenatal diagnosis with chorionic villus sampling (CVS) cells on two women who had experienced neonatal death of SMA type I. DNA extraction was done from archival slide and tissue materials and PEP-PCR was performed using CVS cells. In order to identify common deletion region of SMN and neuronal apoptosis-inhibitory protein (NAIP) genes, cold PCR-SSCP and PCR-restriction site assay were carried out. Case 1 had deletions of the exons 7 and 8, and case 2 had exon 7 only on the telomeric SMN gene. Both cases were found to be normal on NAIP gene. These results were the same for both CVS and archival biopsied specimens. In both cases, the fetuses were, therefore, predicted to be at very high risk of being affected and the pregnancy were terminated. These data clearly demonstrate that archival slide and paraffin-embedded tissues can be a valuable source of DNA when the prenatal genetic diagnosis is needed in case any source for genetic analysis is not readily available due to previous death of the fetus or neonate.

• 생명과학회지
• /
• 제24권9호
• /
• pp.1025-1029
• /
• 2014

자궁경부암은 전세계 여성의 사망원인 2위를 차지하며, 인유두종바이러스 감염과 상관성이 있다. 인유두종바이러스의 백신정책, 병인론, 추적관찰, 역학에서 유전형 검사는 중요하다. 후향적 연구를 위하여 파라핀 조직에서 역교잡반응을 이용하여 인유두종바이러스의 유전형을 검사하는 것은 명확하게 증명된 것은 아니다. 본 연구에서는 자궁경부암 파라핀 조직에서 역교잡반응을 사용하여 인유두종바이러스의 유전형 검사의 유용성을 평가하였다. 총 52개의 자궁경부암 파라핀 조직을 사용하여 역교잡반응을 실시하여 인유두종바이러스의 유전형을 검출하였다. 52개의 파라핀 조직중에서 32(61.5%) 건에서 인유두종바이러스가 검출되었으며, 단순감염이 27(84.4%) 건, 복합감염이 5(15.6%) 건으로 검출되었다. 단순감염에서 인유두종바이러스 유전형은 고위험군 18(8), 58(6), 16(5), 33(1), 35(1), 39(1), 56(1) 유전형과, 저위험군 11(2), 6(1), 70(1) 유전형으로 분석되었다. 복합감염에서 인유두종바이러스 유전형은 16/18(2), 18/52(1), 16/56(1), 16/18/33(1) 유전형으로 검출되었다. 본 연구를 통하여, 파라핀 조직으로 후향적 연구를 위한 인유두종바이러스 유전형 검사가 가능할 것으로 기대된다.

• 대한바이러스학회지
• /
• 제30권1호
• /
• pp.29-37
• /
• 2000

The aim of this study was to investigate viral etiology in dilated cardiomyopathy (DCM) by polymerase chain reaction (PCR) or nested reverse transcription PCR (RT-PCR), and characterize the enteroviral RNA presented in the clinical specimens. Twenty-eight paraffin-embedded heart tissue samples were assayed to detect cytomegalovirus, herpes simplex virus type 1, type 2, parvovirus, adenovirus, and enterovirus (EV) with each specific primer. Of these 28 patients (mean age: 27, M: 24, F: 4), 26 were histologically diagnosed as DCM and 2 as myocardial infarction (MI). Nested RT-PCR detected enteroviral RNA in 7 (26.9%) of 26 patients with DCM, and none of patients with MI. And none of DNA viruses tested were detected from the samples. Amplified products were also genotyped by single-strand conformation polymorphism (SSCP). Three subtypes can be differentiated from 7 clinical specimens. Furthermore, direct sequence analysis was performed to determine whether genetic variation of EV is present in the explanted heart tissues from patients with DCM. Although most of the sequences among the wild isolates have the greatest similarity to those of coxsackievirus B3, there are specific regions of variable sequences (no 490 - no 510). The data suggest that enterovirus may be a major viral pathogen for the DCM in Korea and nucleotide sequence data indicate that coxsackievirus B3 may be a leading etiologic agent of DCM.

• Asian Pacific Journal of Cancer Prevention
• /
• 제13권8호
• /
• pp.3701-3704
• /
• 2012

Background: Cervical cancer is one of the most common cancers in developing countries and the second most common type of cancer in women globally. Several recent studies suggested a co factor role for Chlamydia trachomatis in pathogenesis of cervical cancer. This study aimed to evaluate existence of C. trachomatis DNA in pathologic blocks of patients with cervical cancer. Materials and methods: Seventy-six formaldehyde fixed paraffin embedded tissue specimens from patients with histologically proven history of cervical cancer as well as 150 blocks from healthy peoples were included in the present study. Thin slices were prepared from selected blocks followed by deparaffinization and DNA extraction; the presence of C. trachomatis DNA was examined by Taq Man real-time PCR. Results: Our TaqMan real time PCR assay with cervical specimens of patients with cervical cancer showed that there was no C. trachomatis DNA. Also, we found three positive specimens among our control group. Conclusion: It seems that based on results obtained from the specimens examined in the present study, there is no association between the presence of C. trachomatis DNA in cervical specimens and cervical cancer.

• 대한의생명과학회지
• /
• 제19권2호
• /
• pp.153-157
• /
• 2013

Although culture is the gold standard method to identify mycobacteria, its use in tuberculous lymphadenitis (TBL) is limited due to formalin fixation of the submitted specimens. We evaluated the performance of quantitative real-time PCR (q-PCR) for Mycobacterium Tuberculosis (MTB) in granulomatous lymphadenitis using formalin-fixed paraffin-embedded (FFPE) tissues. From 2000 to 2010, a total number of 117 cases of lymph node samples with granulomatous inflammation which were surgically removed and fixed in formalin were studied. Hematoxylin & Eosin (H&E) and Ziehl-Neelsen-stained (ZN) slides were reviewed. qPCR using Real TB-Taq$^{(R)}$ was performed for all cases to identify Mycobacterium tuberculosis. Thirteen non-tuberculous lymphadenopathy cases were used as negative control. Cervical lymph nodes were more frequently affected (60%, 70/117) than other sites. ZN stain for acid fast bacilli was positive in 19 (16.24%) cases. qPCR for tuberculosis was positive in 92 (78.63%) cases. Caseous necrosis was found in 103 (88.03%) cases. While the ZN stain and qPCR were both negative in all control cases, the qPCR showed a significantly higher positive rate (78.63% vs. 16.24%) compared to ZN stain in histologically diagnosed TBL. Quantitative real-time PCR proves to be more sensitive than ZN stain for diagnosis of tuberculous lymphadenitis.

• Journal of the Korean Association of Oral and Maxillofacial Surgeons
• /
• 제38권3호
• /
• pp.145-151
• /
• 2012

Objectives: Angiogenesis and lymphangiogenesis are correlated with tumor growth and lymph node metastasis in cases of oral squamous cell carcinoma (OSCC). Endoglin is one of the representative vascular endothelial cell markers. Podoplanin is also a representative marker used in order to detect lymphatic endothelial cells. The aim of this study was to determine the correlation between the expression of endoglin/podoplanin and clinical variables associated with OSCC progression. Materials and Methods: Paraffin embedded tissue specimens from 21 patients diagnosed with OSCC were used in this study. Ten patients were diagnosed with early clinical stage (I or II) and 11 patients with advanced clinical stage (III or IV) OSCC. Five patients had positive lymph node involvement. Primary antibodies for endoglin and podoplanin were used to perform the immunohistochemical detection of the vascular and lymphatic endothelial cells. The expression of endoglin and podoplanin was examined by an image analysis program in the three most highly expressed regions of each specimen. Results: The average endoglin expression was observed to be $1.691{\pm}0.920$ in the advanced stage (III, IV) specimens and $0.797{\pm}0.583$ in the early stage (I, II) specimens (P=0.020). The average expression of podoplanin was $0.286{\pm}0.228$ in the advance stage (III, IV) specimens and $0.374{\pm}0.157$ in the early stage (I, II) specimens (P>0.05). There was no statistically significant difference in the expression of endoglin and podoplanin, regardless of whether or not the lymph node was positive. Conclusion: The expression of endoglin was significantly higher in the advanced stage specimens than that in the early stage specimens. Therefore, we concluded that endoglin is a useful molecular marker for use in the evaluation of the progression of OSCC.