• 생명과학회지
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• 제14권2호
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• pp.296-300
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• 2004

환자의 생조직, 얼린 조직 혹은 혈액이 없는 경우에, formalin으로 고정된 파라핀조직을 이용하여 미토콘드리아 돌연변이를 확인할 수 있는지를 조사하였다. MELAS 환자 4명의 파라핀조직을 택해 이들 조직으로부터 DNA를 추출하여 대부분의 MELAS 환자 미토콘드리아 DNA의 tRN $A^{Leu(UUR)}$ gene의 3243지역에서 발견되는 Adenine의 Cuanine으로의 염기치환을 확인하고자 하였다. 실험결과 3명의 환자에게서 이 점 돌연변이를 확인할 수 있어 이들 파라핀조직의 상태가 좋은 것으로 여겨져 미토콘드리아 DNA 돌연변이 연구에 파라핀조직을 활용할 수 있을 것으로 보인다.다.

• 대한의생명과학회지
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• 제28권4호
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• pp.298-306
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• 2022

Pathological tissue fixation using formalin has been widely used for histological samples in many hospitals and institutions. In general, formalin fixatives were either manufactured in laboratories or purchased commercially because of the risks and environmental concerns of handling organic compounds. In this study, the efficacy of three kinds of commercially purchased and one laboratory-made formalin fixative was compared in the PCR-based molecular diagnosis using the extracted DNA from formalin-fixed paraffin-embedded (FFPE) tissues. The quality of extracted DNA from FFPE tonsil tissues with four kinds of formalin solutions was evaluated, and PCR for beta-globin gene and microsatellite instabilities (MSI) tests for pentaplex panel markers were performed using the extracted DNA. There was no difference in PCR and MSI tests as molecular diagnoses regardless of the types of formalin used in this study. However, the total amount and average length of double-stranded DNA extracted from FFPE tonsil tissue showed significant differences according to the type of formalin fixative. Optimized formalin fixatives and methods for DNA extraction might be sophisticated to extract good quality DNA from the small size of specific tissue samples. Further studies are needed to select the most effective formalin fixative for histology and molecular pathology using human FFPE tissues.

• 한국수의병리학회:학술대회논문집
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• 한국수의병리학회 2003년도 추계학술대회초록집
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• pp.27-27
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• 2003

Porcine epidemic diarrhea virus (PEDV) and transmissible gastroenteritis virus (TGEV) infections are considered difficult to distinguish clinically and histopathologically. Prompt differentiation between PEDV- and TGEV-associated enteritis would greatly facilitate the management of disease in countries where PEDV and TGEV are epizootic. Rapid differential diagnosis and treatment are crucial to reducing mortality and morbidity from PEDV- and TGEV-induced enteritis in piglets. The objective for this study was to develop a protocol to differentiate between PEDV and TGEV directly from formalin-fixed, paraffin-embedded tissue, using a multiplex reverse transcription-nested polymerase chain reaction (RT-nPCR) assay. (omitted)

• 대한수의학회지
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• 제33권2호
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• pp.269-275
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• 1993

토끼의 바이러스성 간염, 소위 토끼 출혈병의 원인체에 대한 각종 세포주들에 바이러스증식을 유도하고 한편 실험적 감염예에 대한 면역형광항체법과 immunoperoxidase 방법에 의한 원인체의 조직내에 분포상황을 조사하기 위하여 감염된 토끼를 폐사직후 부검하여 간장, 비장, 신장, 폐 및 뇌조직을 절제하여 동결절편하거나 또는 포르말린고정 파라핀 포매 절편을 $5{\sim}7{\mu}m$로 작제하여 면역반응에 공시하였다. 공시된 각종 세포주에 대한 원인체의 세포배양성은 인정되지 않았으며 면역조직화학적방법을 이용한 면역반응에서는 간장에서 강한 양성반응을 보였으며, 비장과 신장에서는 소수예에서 백색수주변 대식세포와 신사구체에서 양성반응을 각각 나타내었다. 그러나 기타 장기에서는 특이한 양성반응이 인정되지 않았다. ABC immunoperoxidase 방법을 이용한 간장의 포르말린고정 파라핀포매 절편에서 portal triad를 중심으로한 소엽주변부 간세포에서 강한 양성반응을 나타내었으며, 이들 양성반응은 간세포 및 동양혈관세포의 세포질내에 미세과립상으로 미만성 및 세포질주변성으로 관찰되었다. 그리고 감염세포와 비감염세포와의 구별이 명확히 인정되었고 양성반응을 보이는 부위는 H-E 염색상 변성괴사된 간세포에 일치되었다. 이상의 결과에서 본 질병의 표적기관은 간장이며 포르말린고정 파라핀포매 간조직의 immunoperoxidase 방법에 의한 면역조직화학적방법이 본병 진단에 크게 활용되리라 본다.

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

• Asian Pacific Journal of Cancer Prevention
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• 제14권11호
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• pp.6619-6623
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• 2013

Background:The aim of the research was to explore a cost effective, fast, easy to perform, and sensitive method for epidermal growth factor receptor (EGFR) mutation testing. Methods: High resolution melting analysis (HRM) was introduced to evaluate the efficacy of the analysis for dectecting EGFR mutations in exons 18 to 21 using formalin-fixed paraffin-embedded (FFPE) tissues and plasma free DNA from 120 patients. Results: The total EGFR mutation rate was 37.5% (45/120) detected by direct sequencing. There were 48 mutations in 120 FFPE tissues assessed by HRM. For plasma free DNA, the EGFR mutation rate was 25.8% (31/120). The sensitivity of HRM assays in FFPE samples was 100% by HRM. There was a low false-positive mutation rate but a high false-negative rate in plasma free DNA detected by HRM. Conclusions: Our results show that HRM analysis has the advantage of small tumor sample need. HRM applied with plasma free DNA showed a high false-negative rate but a low false-positive rate. Further research into appropriate methods and analysis needs to be performed before HRM for plasma free DNA could be accepted as an option in diagnostic or screening settings.

• 대한의생명과학회지
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• 제19권2호
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• pp.153-157
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• 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.

• 대한의생명과학회지
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• 제8권3호
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• pp.161-165
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• 2002

Apoptosis can be difficult to detect in routine histological sections. Since extensive DNA fragmentation is an important characteristic of this process, visualization of DNA breaks could greatly facilitate the identification of apoptotic cells. Several techniques for the qualitative and quantitative detection of this process have been established; recently, an in situ nick end-labelling technique based on the detection of DNA fragmentation, which is a molecular characteristic of apoptotic cell death, was described. Applying this method to paraffin sections of rat tissues, sensitivity was observed to be inconsistently low with regard to the expected number of apoptotic cells. I describe a new modified method for formalin-fixed, paraffin-embedded tissue sections, pretense pretreatment to permeate the tissue sections that involves an TUNEL (terminal deoxynucleotidyl transferase-mediated dUTP nick end-labeling) is acknowledged as a method of choice in the rapid identification and quantification of the apoptotic cell fraction in paraffin tissue preparations. TUNEL was performed without apoptosis and with apopotosis samples to each of the three concentrations of proteinase K (10, 25, 40 mg/ml) pretreatments. In this study, I show that chemical pretreatments of the tissue sections in proteinase K (25 mg/ml for 15 min at room temperature) considerably enhances the sensitivity of this nick end labelling technique.

• 대한임상검사과학회지
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• 제40권2호
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• pp.113-117
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• 2008

It designed a study to examine the efficiency of DNA and RNA extraction from archival formalin-fixed, paraffin-embedded tissues using an non-heating and heating method. Archival paraffin blocks of liver, kidney, colon were randomly selected. Each paraffin block was prepared in 20 microtubes. For each paraffin blocks were tested non-heating DNA extraction to 10 microtubes and heating protocol under pH 7.0 and $100^{\circ}C$ to 10 microtubes. Evaluation of the results of DNA extraction was carried out by measuring concentration by UV spectrophotometry and then PCR amplification. DNA extraction content that non-heating method was liver $5{\pm}0.7{\mu}g/mL$, kidney $2{\pm}0.3{\mu}g/mL$, colon $6{\pm}0.4{\mu}g/mL$ and heating method was liver $12{\pm}0.6{\mu}g/mL$, kidney $7{\pm}0.5{\mu}g/mL$, colon $10.{\pm}0.3{\mu}g/mL$. Successful RNA extraction was observed, by ${\beta}$-actin amplification, in 46.7% sections for samples treated by the heating method versus 30.0% using non-heating DNA extraction. The extracted nucleic acid showed better values for samples heated at $100^{\circ}C$. Therefore heating extraction of nucleic acid is reliable, quick and efficiency.

• 한국산학기술학회논문지
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• 제20권8호
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• pp.417-424
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• 2019

포르말린을 사용한 조직 고정 방식은 우수한 세포 형태를 유지하며 장기간 조직을 보관할 수 있는 장점이 있으나, 느린 고정 시간, 유해 화학물질에 노출 및 단백질 변형 등의 단점이 있다. 본 연구에서는 마우스의 간과 신장 조직을 이용하여 포르말린 고정과 마이크로파 조사에 의한 빠른 고정을 각각 실시한 후 조직학적 검사와 단백질의 보존 상태를 측정하여 그 결과를 비교하였다. 동일 조직을 절단하여 포르말린 고정과 인산염 완충 식염수에서 마이크로파 조사에 의한 고정 과정을 동시에 실시하였으며, 파라핀 포매 조직에서 제조한 슬라이드에서 H & E와 면역화학염색을 시행하여 조직 고정의 적정성과 항원성을 검사하였다. 또한 고정 조직에서 단백질 추출 양과 질을 각각 BCA법 및 Western blotting법으로 평가하였다. H & E 염색과 면역화학염색을 수행한 결과, 적혈구의 부분적 소실을 제외하고는 마이크로파 고정 조직과 포르말린 고정 조직 간에 대등한 결과를 보였다. 특히, 마이크로파 고정 조직에서 단백질은 잘 보존된 상태로 추출되었다. 결론적으로, 마이크로파 조사를 통한 조직 고정은 포르말린 고정과 비교하여 빠른 고정시간과 우수한 단백질 회수율을 보였으며, 조직 고정의 적정성과 항원성에서도 포르말린 고정과 대등한 결과를 보여, 신속한 조직 고정이 필요한 환경에서 적용이 가능함을 제시하고 있다.