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
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
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
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
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
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
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
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
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
基金the University of Medicine,Pharmacy,Science and Technology Research Grant,No.275/11.01.2017
文摘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.
文摘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.
文摘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.
基金Supported by the Jilin Science & Technology Development Plan,China(No.201201060)the Scientific Research Foundation of Jilin Province,China(No.20100942)+1 种基金the Fund of Developing and Reforming Community of Jilin Province,China(No.2010-1928)the Health Scientific Research Foundation of Jilin Province,China(Nos.2009z081,2010Z083)
文摘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.
文摘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.
文摘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。
文摘本研究以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。
文摘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
文摘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.