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DNA extraction from paraffin embedded colorectal carcinoma samples: A comparison study of manual vs automated methods,using four commercially kits
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作者 Zsolt Kovacs Ioan Jung +4 位作者 Erzsebet Csernak Zoltan Szentirmay Laura Banias Genoveva Rigmanyi Simona Gurzu 《World Journal of Clinical Oncology》 CAS 2019年第9期307-317,共11页
BACKGROUND Nucleic acid isolation from formalin-fixed, paraffin-embedded tissue(FFPET)samples is a daily routine in molecular pathology laboratories, but extraction from FFPET is not always easily achieved. Choosing t... BACKGROUND Nucleic acid isolation from formalin-fixed, paraffin-embedded tissue(FFPET)samples is a daily routine in molecular pathology laboratories, but extraction from FFPET is not always easily achieved. Choosing the right extraction technique is key for further examinations.AIM To compare the performance of four commercially available kits used for DNA extraction in routine practice.METHODS DNA isolation was performed on 46 randomly selected formalin-fixed, paraffinembedded(FFPE) colorectal adenocarcinoma(CRC) surgical specimens. Four commercially available extraction kits were used: two for manual DNA extraction(the Pure Link Genomic DNA Mini Kit from Invitrogen and the High Pure FFPE DNA Isolation Kit from Roche) and two for automated DNA extraction(the i Prep Genomic DNA Kit from Invitrogen and the Magna Pure LC DNA Isolation Kit from Roche). The DNA concentration and quality(odds ratio) among the four systems were compared. The results were correlated with the clinicopathological aspects of CRC cases: age, gender, localization, macro-and microscopic features,lymph node metastases, and the lymph node ratio.RESULTS The highest DNA concentration was obtained using the manual kits: 157.24 ±62.99 ng/μL for the Pure Link Genomic DNA Mini Kit and 86.64 ng/μL± 43.84 for the High Pure FFPE DNA Isolation Kit(P < 0.0001). Lower concentrations were obtained with automated systems: 20.39 ± 21.19 ng/μL for the Magna Pure LC DNA Isolation Kit and 8.722 ± 6.408 ng/μL for the i Prep Genomic DNA Kit,with differences between the systems used(P < 0.0001). The comparison between age, gender, tumor localization, pT or pN stage and the lymph node ratio indicated no statistically significant difference in DNA concentration using any of the nucleic acid isolation kits. DNA concentration was influenced by the macroscopic features and grade of differentiation. A higher DNA concentration was obtained for well-differentiated polypoid colorectal adenocarcinomas(CRCs), compared with undifferentiated ulcero-infiltrative carcinomas,irrespective of the kit used.CONCLUSION For research or diagnosis that needs high DNA concentrations, manual methods of DNA isolation should be used. A higher amount of DNA can be obtained from polypoid-type differentiated CRCs. Automated systems confer comfort and a lower amount of DNA that is, however, sufficient for classic polymerase chain reaction(PCR) and real-time quantitative PCR molecular examinations. All four commercially available kits can be successfully used in daily practice. 展开更多
关键词 DNA ISOLATION Colorectal cancer paraffin-embedDED PureLink GENOMIC DNA Mini KIT High Pure FFPE DNA ISOLATION KIT iPrep GENOMIC DNA KIT MagnaPure LC DNA ISOLATION KIT
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Detection of BCL2-IGHrearrangement on paraffin-embedded tissue sections obtained from a small submucosal tumor of the rectum in a patient with recurrent follicular lymphoma 被引量:3
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作者 Naohisa Yoshida Kenichi Nomura +7 位作者 Yosuke Matsumoto Kazuhiro Nishida Naoki Wakabayashi Hideyuki Konishi Shoji Mitsufuji Keisho Kataoka Takeshi Okanoue Masafumi Taniwaki 《World Journal of Gastroenterology》 SCIE CAS CSCD 2004年第17期2602-2604,共3页
A 59-year-old woman was admitted to our hospital because of recurrent follicular lymphoma(FL).Colonoscopic examination revealed a rectal submucosal tumor(SMT)without any erosions and ulcers.In this patient,it was diff... A 59-year-old woman was admitted to our hospital because of recurrent follicular lymphoma(FL).Colonoscopic examination revealed a rectal submucosal tumor(SMT)without any erosions and ulcers.In this patient,it was difficult to distinguish non-Hodgkin's lymphoma(NHL)invasion from other disorders of the colon including carcinoid tumor merely based on endoscopic findings.Histopathologic and immunohistochemical studies on biopsy specimens showed an infiltration of atypical lymphocytes that were positive for CD20 and BCL2 but negative for UCHL-1.Fluorescence in situ hybridization on paraffin-embedded tissue sections (T-FISH)identified a translocation of BCL2 with IGHgene. Based on these findings,the tumor was defined as an invasion of FL.T-FISH method is useful for the detection of a monoclonality of atypical lymphocytes in an SMT of the gastrointestinal tract,and particularly for the detection of chromosomal translocations specific to lymphoma subtypes. 展开更多
关键词 Gene Rearrangement BIOPSY Female Humans Immunoglobulin Heavy Chains In Situ Hybridization Fluorescence Lymphoma Follicular Middle Aged Neoplasm Recurrence Local paraffin embedding Proto-Oncogene Proteins c-bcl-2 Rectal Neoplasms
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DETECTION OF CYTOMEGALOVIRUS GENOME BY IN SITU HYBRIDIZATION IN PARAFFIN EMBEDDED ENDOMYOCARDIAL BIOPSY SPECIMENS OF VIRAL MYOCARDITIS 被引量:3
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作者 李武 M.Crombach +3 位作者 U.Schonian 杨英珍 B.Maisch 陈灏珠 《Chinese Medical Journal》 SCIE CAS CSCD 1994年第12期10-13,共4页
Cytomegalovirus (CMV) genes were detected by in situ hybridization in 25 Chinese patients with viral myocarditis (VMC). The positive hybridization signals werre found in cardiomyocytes (6 cases, 24%), capillary endoth... Cytomegalovirus (CMV) genes were detected by in situ hybridization in 25 Chinese patients with viral myocarditis (VMC). The positive hybridization signals werre found in cardiomyocytes (6 cases, 24%), capillary endothelial cells (4 cases, 16%) and interstitial cells (7 cases, 28%). The difference between VMC and control group (16 cases died of brain trauma and 10 cases of congenital heart diseases was statistically significant. There was no definite pathomorphological relationship between the detection of CMV genes and myocardial lesions. The results suggest that CMV infection may be one of the causes of myocarditis and chronic stimulation of the immune system induced by CMV may be a possible pathogenesis of this disease. 展开更多
关键词 VMC DETECTION OF CYTOMEGALOVIRUS GENOME BY IN SITU HYBRIDIZATION IN paraffin embedDED ENDOMYOCARDIAL BIOPSY SPECIMENS OF VIRAL MYOCARDITIS EMB In
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DNA Extraction from Formalin-fixed and Paraffin-embedded Tissues by Triton X-100 for Effective Amplification of EGFR Gene by Polymerase Chain Reaction 被引量:1
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作者 WANG Xiao-feng DU Zhen-wu +3 位作者 WU Meil ZHANG Yu-cheng JIANG Yang ZHANG Gui-zhen 《Chemical Research in Chinese Universities》 SCIE CAS CSCD 2012年第4期662-665,共4页
For first-line non-small-cell lung cancer(NSCLC) therapy,detecting mutation status of the epidermal growth factor receptor(EGFR) gene constitutes a prudent test to identify patients who are most likely to benefit ... For first-line non-small-cell lung cancer(NSCLC) therapy,detecting mutation status of the epidermal growth factor receptor(EGFR) gene constitutes a prudent test to identify patients who are most likely to benefit from EGFR-tyrosine kinase inhibitor(TKI) therapy.Now,the material for detecting EGFR gene mutation status mainly comes from formalin-fixed and paraffin-embedded(FFPE) tissues.DNA extraction from FFPE and the amplification of EGFR gene by polymerase chain reaction(PCR) are two key steps for detecting EGFR gene mutation.We showed a simple method of DNA extraction from FFPE tissues for the effective amplification of EGFR gene.Extracting DNA from the FFPE tissues of NSCLC patients with 1% Triton X-100(pH=10.0) was performed by heating at 95 °C for 30 min.Meanwhile,a commercial kit was used to extract DNA from the same FFPE tissues of NSCLC patients for comparison.DNA extracted products were used as template for amplifying the exons 18,19,20 and 21 of EGFR by PCR for different amplified fragments.Results show that DNA fragment size extracted from FFPE tissues with 1% Triton X was about 250―500 base pairs(bp).However,DNA fragment size extracted from FFPE tissues via commercial kit was about from several hundreds to several thousands bp.The DNA yield extracted from FFPE tissues with 1% Triton X was larger than that via commercial kit.For about 500 bp fragment,four exons of EGFR could not be amplified more efficiently from extracted DNA with 1% Triton X than with commercial kit.However,for about 200 bp fragment.This simple and non-laborious protocol could successfully be used to extract DNA from FFPE tissue for the amplification of EGFR gene by PCR,further screening of EGFR gene mutation and facilitating the molecular analysis of a large number of FFPE tissues from NSCLC patients. 展开更多
关键词 EGFR gene amplification DNA extraction Formalin-fixed and paraffin-embedded tissue Non-small-cell lung cancer
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Trefoil Factor 3 (TFF3) mRNA Expression Level in Follicular Thyroid Tumors Using Formalin-Fixed, Paraffin-Embedded (FFPE) Blocks
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作者 Saydiganikhodja Ismailov Murodjon Rashitov +5 位作者 Makio Kobayashi Noriyuki Shibata Yoichiro Kato Yoko Omi Masatoshi Iihara Takahiro Okamoto 《Open Journal of Pathology》 2013年第2期78-84,共7页
Background: Differential diagnosis of follicular thyroid carcinoma (FTC) from follicular thyroid adenoma (FTA) is often difficult since presence or absence of capsular/vascular invasion can not be determined by preope... Background: Differential diagnosis of follicular thyroid carcinoma (FTC) from follicular thyroid adenoma (FTA) is often difficult since presence or absence of capsular/vascular invasion can not be determined by preoperative fine needle aspiration cytology, and may not be judged unanimously on permanent sections even among experienced pathologists. Determination of molecular-genetic factors such as trefoil factor 3 (TFF3) mRNA in the follicular thyroid tumors may be useful aid to improve the accuracy of diagnosis, though it is considered to be unstable and relatively low concentrated genetic substance. Purpose of our study is to investigate expression level of TFF3 mRNA of thyroid follicular tumors using formalin-fixed, paraffin-embedded (FFPE) tissue. Methods: Study population included FFPE sections from 19 FTC cases, 20 FTA cases, 11 adenomatous goiter (G) cases and 12 samples of normal thyroid tissue (N) adjacent to thyroid tumors. RNeasy FFPE kit was used for extraction of total RNA. Purification and concentration values were determined by spectrophotometer. Extracted RNA was used for cDNA synthesis in reverse transcription. Synthesized cDNA subsequently proceeded for relative quantification of TFF3 mRNA by RT-qPCR using TFF3 primers. Glyceroldehyde-3-phosphate dehydrogenase (GAPDH) and hypoxanthin phosphorobosyltransferase1 (HPRT1) were used as control genes. The mean and standard deviation of TFF3 mRNA expression level were analyzed by software Multiplate RQ. Results: Extraction by the FFPE kit yielded high concentration of RNA in all cases. Purification values were 1.8 in average. Concentration values were significantly higher in FTC and FTA relative to G and N tissues, possibly due to high density of thyrocytes in the samples. Relative quantification of TFF3 mRNA expression level showed broad ranges both in FTC and FTA, while the analyses in G and N tissues indicated narrow ranges. Conclusion: FFPE tissues from thyroid follicular tumors can be used for measurement of unstable and low concentrated genetic substances such as TFF3 mRNA. Its diagnostic value yet remains to be determined. 展开更多
关键词 FOLLICULAR Thyroid Tumors TFF3 RT-qPCR FORMALIN-FIXED paraffin-embedDED Tissue
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围绕FFPE样本特性的DNA纯化纳米磁珠优化
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作者 肖林 吕晓芳 +4 位作者 曹师瑜 江丹 孙林雍 李坤 叶丰 《临床与实验病理学杂志》 CAS 北大核心 2024年第7期725-730,共6页
目的 探讨围绕福尔马林固定石蜡包埋(formalin fixed paraffin embedded, FFPE)样本特性获取更高质量/得率的纳米磁珠核酸提取方案,改进分子病理技术。方法 合成4大类15个小类的备选磁珠,以FFPE样本为中心,筛选高质量/得率磁珠。模拟常... 目的 探讨围绕福尔马林固定石蜡包埋(formalin fixed paraffin embedded, FFPE)样本特性获取更高质量/得率的纳米磁珠核酸提取方案,改进分子病理技术。方法 合成4大类15个小类的备选磁珠,以FFPE样本为中心,筛选高质量/得率磁珠。模拟常规组织、粗针穿刺(肝脏)、纤维支气管镜样本(肺),装管相同张数连片。采用筛选的最佳磁珠与市场出售的常见磁珠试剂提取核酸,横向对比纯化总量、片段大小等质量参数。应用PCR和Sanger验证核酸的下游应用。结果 以FFPE样本DNA为中心筛选自制纳米磁珠,获得最佳性能纳米磁珠总回收率为58.5%±1.58%,5种市售商品化磁珠和3种国产磁珠总回收率为18.68%~40.71%。相同组织量(连片)提取的DNA总量,在模拟常规组织、粗针穿刺和纤维支气管镜样本核酸得率比市售试剂盒提高39.49%~181.72%(P<0.05)。模拟纤维支气管镜样本1张(4μm)总量可达100 ng以上、5张总量可达400 ng以上。结论 以FFPE样本DNA为中心筛选的DNA纯化纳米磁珠,与商品化磁珠相比有较大提升,为临床分子病理检测的质量保证、自动化检测和项目拓展提供空间。 展开更多
关键词 石蜡包埋组织 DNA提取 纳米磁珠 PCR Sanger
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石蜡包埋机评价指标体系的建立与应用 被引量:1
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作者 卢如意 孙静 +2 位作者 王之晨 张倩 冯靖祎 《中国医院建筑与装备》 2024年第1期84-88,共5页
采用文献回顾、专家访谈、小组讨论等方法拟定石蜡包埋机评价指标体系草案及专家咨询问卷,采用德尔菲法确定评价指标,应用层次分析法构建判断矩阵得到评价指标权重,通过层次单排序及层次总排序一致性检验,形成石蜡包埋机评价指标体系,... 采用文献回顾、专家访谈、小组讨论等方法拟定石蜡包埋机评价指标体系草案及专家咨询问卷,采用德尔菲法确定评价指标,应用层次分析法构建判断矩阵得到评价指标权重,通过层次单排序及层次总排序一致性检验,形成石蜡包埋机评价指标体系,最后邀请临床医务人员根据已建立的评价指标体系开展石蜡包埋机评价。构建了一套石蜡包埋机评价指标体系,包含5个一级指标、26个二级指标,专家权威程度在满意范围之内;临床医务人员对国产某品牌石蜡包埋机进行试用并评价,从冷冻台和包埋机两个方面共计提出了18条改进意见建议。本研究所建立的石蜡包埋机评价指标体系,专家参与度高、积极性好、权威程度高,说明结果可参考。 展开更多
关键词 石蜡包埋机 评价指标体系 权重 德尔菲法 层次分析法
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胸腹水液基细胞学检测联合胸腹水沉淀物琼脂石蜡双包埋切片法在病理检测的价值
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作者 田莉 吴阳春 《中华养生保健》 2024年第3期81-84,共4页
目的探讨与分析胸腹水液基细胞学检测联合胸腹水沉淀物琼脂石蜡双包埋切片法在病理检测的价值。方法选取2019年3月—2022年9月在乌鲁木齐市中医医院诊治的92例胸腹水患者作为研究对象,所有患者都给予胸腹水液基细胞学检测联合胸腹水沉... 目的探讨与分析胸腹水液基细胞学检测联合胸腹水沉淀物琼脂石蜡双包埋切片法在病理检测的价值。方法选取2019年3月—2022年9月在乌鲁木齐市中医医院诊治的92例胸腹水患者作为研究对象,所有患者都给予胸腹水液基细胞学检测联合胸腹水沉淀物琼脂石蜡双包埋切片法,都给予病理活检,判断诊断的价值。结果在92例患者中,胸腹水液基细胞学检测判断为恶性肿瘤54例,良性肿瘤38例,胸腹水液基细胞学检测诊断恶性肿瘤的敏感度与特异度分别为85.48%、96.67%。胸腹水沉淀物琼脂石蜡双包埋切片检测判断为恶性肿瘤55例,良性肿瘤37例,胸腹水沉淀物琼脂石蜡双包埋切片检测诊断恶性肿瘤的敏感性与特异性分别为88.71%、100.00%。胸腹水液基细胞学检测联合胸腹水沉淀物琼脂石蜡双包埋切片检测判断为恶性肿瘤61例,良性肿瘤31例,胸腹水液基细胞学检测联合胸腹水沉淀物琼脂石蜡双包埋切片检测诊断恶性肿瘤的敏感度与特异度分别为98.39%、100.00%。恶性肿瘤患者的人表皮生长因子受体2、突触素、嗜铬素A表达阳性率分别为40.32%、41.94%、43.55%,与良性肿瘤患者的10.00%、13.33%、10.00%相比有显著提高,差异有统计学意义(P<0.05)。胸腹水液基细胞学检测联合胸腹水沉淀物琼脂石蜡双包埋切片法对肿瘤良恶性的诊断敏感性高于单项检测,差异有统计学意义(P<0.05),不同检测方法的诊断特异性比较,差异无统计学意义(P>0.05)。ROC曲线分析显示,胸腹水液基细胞学检测联合胸腹水沉淀物琼脂石蜡双包埋切片法对肿瘤良恶性的诊断曲线下面积为0.883。结论胸腹水液基细胞学检测联合胸腹水沉淀物琼脂石蜡双包埋切片法在病理检测中的应用能提高诊断敏感度,还可保持非常高的诊断特异度,对患者的原发肿瘤良恶性判定具有很好的价值。 展开更多
关键词 胸腹水 液基细胞学 胸腹水沉淀物 琼脂石蜡双包埋切片 恶性肿瘤 良性肿瘤
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插入缺失位点和miniSTR在甲醛固定石蜡包埋组织中的应用检测
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作者 王亚丽 陈丽琴 +7 位作者 王嘉栎 熊磊 孔倩倩 白志美 赵阳 白慧茹 顾捷 宋振祥 《解剖学杂志》 CAS 2023年第4期317-323,共7页
目的:研究插入缺失位点InDel与mini短串联重复序列(STR)遗传标记对甲醛固定石蜡包埋组织应用的价值。方法:19份甲醛固定石蜡包埋组织作为疑难检材,应用3种常染色体遗传标记多重扩增系统,即InDel系统(Investigatior■DIPplex Kit)、mini... 目的:研究插入缺失位点InDel与mini短串联重复序列(STR)遗传标记对甲醛固定石蜡包埋组织应用的价值。方法:19份甲醛固定石蜡包埋组织作为疑难检材,应用3种常染色体遗传标记多重扩增系统,即InDel系统(Investigatior■DIPplex Kit)、miniSTR系统(华夏白金扩增试剂盒)和STR系统(Goldeneye®20A),对STR分型不完整或失败的检材,再分别用InDel和miniSTR系统进行DNA分型检测,比较两者的分型质量和检出率。结果:19份甲醛固定石蜡包埋组织经Goldeneye 20A试剂盒检测,均分型不完整。应用miniSTR系统检测显示,7份样本的检出率100%,其中,D19S433基因座的检出率为100%,D5S818、D13S817和D1S1656基因座的检出率最小,为36.84%。应用InDel系统检测发现,1份样本InDel位点电泳分型完整,所有样本的位点丢失率均小于50%,其中7份样本的位点丢失率低至10%以下。结论:应用遗传标记对甲醛固定石蜡包埋组织这类疑难降解检材进行检测,插入缺失多态性位点InDel与miniSTR可以作为联合检测标记。 展开更多
关键词 插入/缺失多态性位点 短串联重复序列 甲醛固定石蜡包埋 降解检材
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二代测序建库中福尔马林固定石蜡包埋标本DNA超声打断方法的优化效果观察
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作者 李余发 许洁 《医药前沿》 2023年第20期24-27,共4页
目的:分析24例福尔马林固定石蜡包埋(FFPE)标本DNA采用新的改良的超声打断方法得到的结果与原打断方法得到结果的一致性,进一步优化目前福尔马林固定石蜡包埋样本DNA超声打断方法。方法:收集广东省人民医院病理科2023年4—5月进行二代... 目的:分析24例福尔马林固定石蜡包埋(FFPE)标本DNA采用新的改良的超声打断方法得到的结果与原打断方法得到结果的一致性,进一步优化目前福尔马林固定石蜡包埋样本DNA超声打断方法。方法:收集广东省人民医院病理科2023年4—5月进行二代测序检测病例24例,建库过程中根据使用的打断方法不同分为两组,1组采用原DNA打断方法(打断条件为:稀释后样本转移入配套的microTUBE AFA Fiber Snap-Cap打断管),2组采用改良DNA打断方法(打断条件为:稀释后样本转移入一次性超声波核酸打断耗材),比较两组实验结果是否一致。结果:两组实验结果得到的插入片段大小均>150bp,符合二代测序实验需要,差异无统计学意义(P>0.05)。结论:在临床实践中,二代测序建库过程中采用新的福尔马林固定石蜡包埋标本DNA超声打断方法可以达到原方法的实验效果,且更加经济实惠,从而能降低二代测序实验成本,值得在临床应用。 展开更多
关键词 DNA 超声打断 福尔马林固定石蜡包埋 二代测序
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穿刺小标本改良石蜡块制作方法及评价 被引量:2
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作者 陈宛灵 石超吉 +2 位作者 王敏 顾挺 田臻 《上海口腔医学》 CAS 北大核心 2023年第1期6-11,共6页
目的 :改良现有琼脂预包埋石蜡包埋法,评价该包埋法对空芯针穿刺活检小标本组织完整性、组织学形态、蛋白质和DNA检测的影响。方法:取10例口腔黏膜鳞癌空芯针穿刺活检标本,分别进行成型包埋模具的改良琼脂预包埋石蜡包埋法及常规石蜡包... 目的 :改良现有琼脂预包埋石蜡包埋法,评价该包埋法对空芯针穿刺活检小标本组织完整性、组织学形态、蛋白质和DNA检测的影响。方法:取10例口腔黏膜鳞癌空芯针穿刺活检标本,分别进行成型包埋模具的改良琼脂预包埋石蜡包埋法及常规石蜡包埋法,前者缩短脱水时间至3.5 h,后者为12 h。样本处理后,分别行H-E染色、组织学形态、免疫组织化学(IHC)和DNA荧光原位杂交(FISH)等检测,并对检测结果进行对比分析。采用GraphPad Prism 9软件包对数据进行组间t检验。结果:改良琼脂预包埋法较琼脂预包埋法降低了操作难度,易于推广。与常规石蜡包埋法相比,组织脱水时间显著缩短(P<0.001),对镜下组织学形态及后续IHC和FISH检测无影响。结论:改良琼脂预包埋石蜡包埋法满足临床病理诊断对组织处理的要求,是值得推广的空芯针穿刺活检标本组织包埋法。 展开更多
关键词 空芯针穿刺活检 琼脂预包埋 石蜡包埋
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实时荧光定量PCR在结核病理诊断中的应用
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作者 袁风菊 王宪伟 +3 位作者 徐培 成思颖 李慧 丁兀兀 《诊断病理学杂志》 2023年第6期541-544,548,共5页
目的探讨实时荧光定量PCR(qPCR)在结核病理诊断中的应用价值。方法回顾性分析1323例疑似结核病的石蜡包埋组织qPCR结果,并与临床诊断结果、抗酸染色结果进行对比分析。结果1323例病例中1073例有明确的诊断结果,其中结核阳性率为68.22%(7... 目的探讨实时荧光定量PCR(qPCR)在结核病理诊断中的应用价值。方法回顾性分析1323例疑似结核病的石蜡包埋组织qPCR结果,并与临床诊断结果、抗酸染色结果进行对比分析。结果1323例病例中1073例有明确的诊断结果,其中结核阳性率为68.22%(732/1073)。1073例病例中,qPCR阳性率为45.39%、敏感性为0.663、特异性为0.994,肺、胸膜、骨、淋巴结、肠中qPCR敏感性分别为0.706、0.676、0.437、0.678、0.625,特异性分别为0.993、1.000、1.000、1.000、1.000,手术组织阳性率和敏感性分别为48.42%、0.692,均高于活检组织,且差异有统计学意义(P<0.05)。755例同时具有qPCR结果和抗酸染色结果的病例中,qPCR与抗酸染色法符合率75.36%(95%CI 72.17~78.30),qPCR阳性率(43.44%,328/755)高于抗酸染色的阳性率(21.5%,162/755)(P<0.05),qPCR的AUC(95%CI)=0.837(0.817~0.859)高于抗酸染色AUC(95%CI)=0.656(0.634~0.678)(P<0.05),敏感性高于抗酸染色(0.677 vs 0.327)(P<0.05)。结论qPCR在结核病的病理诊断中具有良好的应用价值,联合抗酸染色可以更准确地进行鉴别诊断。 展开更多
关键词 结核分枝杆菌 石蜡包埋组织 荧光定量PCR 抗酸染色
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甲醛固定石蜡包埋组织4种DNA提取方法的比较分析
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作者 黄静 李琪睿 +11 位作者 李禹泽 宋海迪 刘安琪 宋振祥 赵晋远 宣宇佳 白慧茹 霍志鑫 易新容 顾捷 王亚丽 陈丽琴 《激光生物学报》 CAS 2023年第5期441-449,共9页
本研究以10例甲醛固定石蜡包埋组织(FFPET)作为疑难检材,将其制成蜡膜,分别以二甲苯-Chelex100法、脱蜡液-Chelex100法、脱蜡液-Vazyme FastPure FFPE DNA Isolation试剂盒(简称FP试剂盒)、脱蜡液-北京天根石蜡包埋组织基因组提取试剂盒... 本研究以10例甲醛固定石蜡包埋组织(FFPET)作为疑难检材,将其制成蜡膜,分别以二甲苯-Chelex100法、脱蜡液-Chelex100法、脱蜡液-Vazyme FastPure FFPE DNA Isolation试剂盒(简称FP试剂盒)、脱蜡液-北京天根石蜡包埋组织基因组提取试剂盒法(简称天根试剂盒)提取DNA,进行紫外分光光度计测定(浓度、纯度)及短串联重复序列(STR)分型检测(23个常染色体),通过比较分析FFPET在4种不同提取方法中的提取质量、STR分型质量和检出率,从而寻求一种高效简便、经济实用、方便快捷的甲醛固定石蜡包埋组织中DNA提取方法。结果表明:应用4种方法提取的石蜡包埋组织蜡膜DNA经紫外分光光度计测定后显示,应用2种Chelex100法提取的DNA浓度高于2款试剂盒,试剂盒提取的DNA的纯度优于另外2种方法,以上差异均具有统计学意义(P<0.05);FP试剂盒提取DNA的平均浓度和纯度均优于天根试剂盒,无显著性差异(P>0.05);STR分型普遍存在不同基因座和同一基因座不同等位基因之间的扩增不均衡,呈现前高后低状态;STR平均检出率为FP试剂盒>天根试剂盒>Chelex100法,2种Chelex100法STR基因座检出率比较无显著差异(P>0.05),但分别和另外2种方法试剂盒方法相比有显著差异(P<0.05),FP试剂盒和天根试剂盒法相比有显著差异(P<0.05)。因此,在涉及甲醛固定石蜡包埋组织这类疑难检材的法医DNA鉴定案件时,脱蜡液-Vazyme FastPure FFPE DNA Isolation试剂盒法优于其余3种提取DNA方法。 展开更多
关键词 甲醛固定石蜡包埋组织 法医检材 疑难检材 方法比较
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Detection of SYT-SSX fusion transcripts in paraffin-embedded tissues of synovial sarcoma by reverse transcription-polymerase chain reaction 被引量:2
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作者 魏永昆 王坚 +3 位作者 朱雄增 施达仁 久冈正典 桥本洋 《Chinese Medical Journal》 SCIE CAS CSCD 2002年第7期1043-1047,151,共5页
OBJECTIVE: To assess the feasibility of detecting SYT-SSX fusion transcripts in paraffin-embedded tissues of synovial sarcoma by reverse transcription-polymerase chain reaction (RT-PCR). METHODS: RT-PCR was used to am... OBJECTIVE: To assess the feasibility of detecting SYT-SSX fusion transcripts in paraffin-embedded tissues of synovial sarcoma by reverse transcription-polymerase chain reaction (RT-PCR). METHODS: RT-PCR was used to amplify the SYT-SSX fusion transcripts using archival formalin-fixed paraffin-embedded tumor specimens from a series of 37 synovial sarcoma cases. To investigate the specificity of the SYT-SSX fusion transcripts, a variety of non-synovial sarcoma tumors were included in the study as negative controls. The detected messages derived from fusion genes were confirmed by subsequent sequence analysis. RESULTS: SYT-SSX fusion transcripts were detected in 33 of 37 (89.2%) synovial sarcomas. None of the 34 cases of non-synovial sarcoma tumors showed amplified products of SYT-SSX fusion transcripts, although PBGD mRNA was detected in all specimens. Among 33 SYT-SSX-positive synovial sarcomas, 22 tumors had an SYT-SSX 1 fusion transcript, whereas 6 tumors had an SYT-SSX2 fusion transcript. Fusion types can not be distinguished in the remaining 5 cases. There was a significant relationship between SYT-SSX fusion type and histologic subtype. All 10 biphasic synovial sarcomas had the SYT-SSX1 fusion, whereas all tumors with SYT-SSX2 were of monophasic morphology (P 展开更多
关键词 Reverse Transcriptase Polymerase Chain Reaction ADOLESCENT ADULT Aged Aged 80 and over Female Humans Male Middle Aged Oncogene Proteins Fusion paraffin embedding RNA Messenger Sarcoma Synovial
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Tectona grandis (Teak Tree) Young Leaf Extract as a Histological Stain
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作者 Cecilia Smith-Togobo Adam Abdul Fatau +4 位作者 Magalys Cuba Lopez Felix Kpor David Larbi Simpong George Osei Yiadom Emmanuel Akomanin Asiamah 《Journal of Biomedical Science and Engineering》 CAS 2023年第2期17-41,共25页
Stains are applied to impart contrast to the tissue and identify particular features of interest. However, the use of synthetic dyes as staining reagents has been associated with significant human health challenges an... Stains are applied to impart contrast to the tissue and identify particular features of interest. However, the use of synthetic dyes as staining reagents has been associated with significant human health challenges and pollution of the ecosystem. These developments have necessitated a shift towards using natural dyes that are eco-friendlier and readily available. We investigated the staining reaction patterns of teak tree leaves (Tectona grandis) dye extracts and explored their suitability as a cytoplasmic stain in micromorphological assessments. Dye extracts were prepared using acetone, methanol, and ethanol as solvents from air-dried (under shade) teak tree young leaves. The dye extracts were applied as a counterstain and evaluated against eosin in formalin-fixed paraffin-embedded (FFPE) bovine tissue sections at varying concentrations and different staining times. Teak tree leaves (Tectona grandis) dye extracts produced relatively varying staining intensities of reddish-brown cytoplasmic coloration when used on bovine tissue at different concentrations and staining times comparable to eosin and with blue-purple hematoxylin nuclear stain. The present study showed that Tectona grandis leaf dye extracts provide an excellent cytoplasmic staining pattern and can be used as an alternative counterstain in routine H&E staining techniques. 展开更多
关键词 Histology CYTOPLASM Plant Extract Tectona grandis Leaves Formalin-Fixed paraffin-embedded Tissues Natural Dye STAINING Cytoplasmic Stain Animal Tissues Staining Reaction
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琼脂石蜡双包埋法制备斑马鱼幼鱼切片
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作者 张颖 夏铭 +1 位作者 黄春华 楼迪栋 《安徽农学通报》 2023年第21期80-84,共5页
为优化探索斑马鱼幼鱼石蜡组织切片制作方法,以发育4d的斑马鱼幼鱼为试验材料,经4%多聚甲醛固定24h后进行琼脂预包埋、脱水、透明、浸蜡再包埋切片进行苏木精-伊红(HE)染色,比较琼脂-石蜡双包埋法与普通石蜡包埋法之间的优势。结果表明... 为优化探索斑马鱼幼鱼石蜡组织切片制作方法,以发育4d的斑马鱼幼鱼为试验材料,经4%多聚甲醛固定24h后进行琼脂预包埋、脱水、透明、浸蜡再包埋切片进行苏木精-伊红(HE)染色,比较琼脂-石蜡双包埋法与普通石蜡包埋法之间的优势。结果表明,琼脂石蜡双包埋法能保持斑马鱼幼鱼的形态结构,组织无变形、碎裂,镜下结果明显比普通石蜡包埋法清楚;HE染色效果好。琼脂石蜡双包埋法可以克服普通石蜡包埋法的不足,设备简单,操作简便,易于普及推广。 展开更多
关键词 斑马鱼 琼脂 石蜡 双包埋法 石蜡切片
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比较胸腹水液基细胞学剩余标本制作细胞块的3种方法 被引量:12
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作者 王守梅 杨秋红 +2 位作者 魏静静 王雨稼 张树辉 《第二军医大学学报》 CAS CSCD 北大核心 2013年第2期160-163,共4页
目的优化胸腹水液基细胞学剩余标本制作细胞块的程序,并探讨其在病理诊断中的应用价值。方法 150例胸腹水薄层液基细胞学技术(thinprep cytologic test,TCT)检测剩余标本,根据细胞学诊断结果分为3组,每组50例,分别采用直接离心法、蛋清... 目的优化胸腹水液基细胞学剩余标本制作细胞块的程序,并探讨其在病理诊断中的应用价值。方法 150例胸腹水薄层液基细胞学技术(thinprep cytologic test,TCT)检测剩余标本,根据细胞学诊断结果分为3组,每组50例,分别采用直接离心法、蛋清液作为支架法和细胞块试剂盒法处理后制作石蜡细胞块,对比分析3种不同方法制成的细胞切片上的恶性细胞检出率、细胞分布状况和形态特征,并初步比较细胞块和组织块免疫化学染色效果。结果 150例胸腹水TCT检测出的恶性细胞率为31.3%(47/150),其剩余标本经细胞块法检测出的总恶性细胞检出率为40.7%(61/150),其中直接离心法、蛋清液作为支架法和细胞块试剂盒法的检出率分别为26.0%(13/50)、46.0%(23/50)和50.0%(25/50)。用蛋清液作为支架及细胞块试剂盒两种方法制作的细胞块,恶性细胞检出率高于直接离心法(P<0.05),细胞聚集度和细胞分布也优于直接离心法,细胞块和组织块免疫化学染色效果相似。结论用蛋清液作为支架及细胞块试剂盒两种方法制作的细胞块可提高胸腹水液基细胞学剩余标本的恶性细胞检出率,并可用于免疫细胞化学检测。 展开更多
关键词 胸腔积液 腹水 液基细胞学 细胞块 石蜡包埋
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肺癌石蜡包埋组织及新鲜标本组织芯片制备方法学探讨 被引量:17
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作者 韩瑞刚 张建中 +4 位作者 黄英武 郑燕华 赵尔增 周金莲 张卫国 《临床与实验病理学杂志》 CAS CSCD 2002年第4期404-407,共4页
目的 :探讨和建立石蜡包埋组织和新鲜冷冻标本制备组织芯片的方法 ,以及在基因表达分析中的应用。方法 :选取肺癌石蜡包埋组织 90例 ,新鲜液氮冻存肺癌及癌旁肺组织 2 0例 ,常规切片HE染色下确定穿刺部位 ,石蜡包埋组织芯片制作按Konone... 目的 :探讨和建立石蜡包埋组织和新鲜冷冻标本制备组织芯片的方法 ,以及在基因表达分析中的应用。方法 :选取肺癌石蜡包埋组织 90例 ,新鲜液氮冻存肺癌及癌旁肺组织 2 0例 ,常规切片HE染色下确定穿刺部位 ,石蜡包埋组织芯片制作按Kononem等方法 (1998) ,每例穿取 0 6mm组织柱 2条 ,制备 180点阵组织芯片 ;而新鲜组织芯片参考Fejzo等方法 (2 0 0 1) ,并略作改良 ,制备 5 0点阵组织芯片。所有组织芯片模块 ,经 4 μm切片、常规组织学染色和免疫组化染色 ,以评价其在形态学及基因表达分析中应用的可行性。结果 :180点阵石蜡包埋肺癌组织芯片 ,组织排列整齐 ,HE染色后组织形态可观察率在 98%以上 ;p5 3、c myc、bcl 2、bax和Ki 6 7等免疫组化染色后 ,肺癌组织可见不同程度的抗原表达 ,组织点阵的形态可观测率在 89%~ 95 %之间。新鲜组织芯片经HE染色后 ,形态可观测率和基因表达可分析率在 70 %左右。结论 :手工法组织芯片制备简便易行 ,使用组织少 ,不破坏原蜡块 ;实验均一性、可比性好 ,且节省试剂。与常规组织切片相比 ,2点 0 6mm组织可以获得原蜡块 95 %以上的信息 ,完全可以用于组织形态学和蛋白水平上的基因表达分析 ;新鲜组织芯片技术已经初步建立 ,但尚有待于进一步完善制备技术和mRNA分析方法。 展开更多
关键词 肺癌 石蜡包埋组织 新鲜标本组织芯片 制备 方法学
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普通甲醛固定石蜡包埋组织DNA提取方法的探讨 被引量:18
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作者 田子强 刘俊峰 +3 位作者 张少为 李保庆 王福顺 张月峰 《癌症》 SCIE CAS CSCD 北大核心 2004年第3期342-345,共4页
背景与目的:在国内,几乎所有的医院和科研机构均应用普通甲醛固定手术标本,但因为普通甲醛固定石蜡包埋组织中的DNA降解相对严重,从这些蜡块中提取高质量的DNA非常困难。本研究的目的是寻找从普通甲醛固定石蜡包埋组织中提取DNA的理想... 背景与目的:在国内,几乎所有的医院和科研机构均应用普通甲醛固定手术标本,但因为普通甲醛固定石蜡包埋组织中的DNA降解相对严重,从这些蜡块中提取高质量的DNA非常困难。本研究的目的是寻找从普通甲醛固定石蜡包埋组织中提取DNA的理想方法。方法:取我院2000年甲醛固定石蜡包埋的手术切除食管癌标本15例,分别以蛋白酶K消化和不同pH值下加热两种裂解方法裂解细胞,以酚氯仿抽提法提取其DNA;并在蛋白酶K消化法进行DNA提取的过程中,应用交叉设计对二戊烯或二甲苯脱蜡、37℃或56℃消化48h或72h、氯化钠盐析或酚氯仿抽提4种参数进行研究,所得DNA质量以电泳分析和PCR扩增结果判断。结果:蛋白酶K消化酚氯仿抽提法所得DNA产量(平均值为17.88μg)和质量均高于pH值为7~12条件下的加热提取法(P<0.05)。56℃消化所得的DNA质量明显优于37℃,而消化72h的DNA亦明显优于48h者。不同脱蜡方法和抽提方法对DNA的产量和质量影响不大,结论:以蛋白酶K消化氯化钠盐析法,从普通甲醛固定石蜡包埋组织中提取DNA质量较可靠。并且,应用蛋白酶K于56℃消化3天更能获得高质量和高产量的DNA。 展开更多
关键词 甲醛 石蜡包埋组织 DNA提取方法 食管肿瘤
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甲醛固定石蜡包埋组织STR分型检测 被引量:10
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作者 柳燕 李莉 +2 位作者 赵珍敏 张素华 赵书民 《法医学杂志》 CAS CSCD 2009年第5期337-340,344,共5页
目的评估10%甲醛固定石蜡包埋组织STR分型结果的影响因素。方法采用QIAGEN法、IQ法、Chelex法对2具新鲜尸体在尸检时常规制备的心、脑、肝、脾、肾、肺、胃、肠石蜡包埋组织进行DNA提取,用AmpFlSTR Identifiler试剂盒进行PCR扩增,在3100... 目的评估10%甲醛固定石蜡包埋组织STR分型结果的影响因素。方法采用QIAGEN法、IQ法、Chelex法对2具新鲜尸体在尸检时常规制备的心、脑、肝、脾、肾、肺、胃、肠石蜡包埋组织进行DNA提取,用AmpFlSTR Identifiler试剂盒进行PCR扩增,在3100-Avant上完成片段分析。另外对15个案例中室温保存1~5年的心、肝、肺、肠存档石蜡包埋组织共56份采用同样的方法进行STR分型。以STR基因座检出率评估分型有效性。结果各种组织DNA片段均随着保存时间延长而持续降解,其中心、肺组织STR基因座检出率与保存时间存在线性相关。相同保存时间时,各种组织基因座检出率差异有统计学意义。其中以肺组织在不同保存时间中基因座检出率最高。结论在甲醛固定时间一定的条件下,存放时间、组织类型、DNA提取方法和PCR模板质量浓度是影响石蜡包埋组织STR分型的重要因素。 展开更多
关键词 法医遗传学 石蜡包埋 STR分型 影响因素
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