肾脏移植物血管内皮细胞嵌合现象预后观察

目的:用荧光原位杂交方法(FISH)观察移植肾中血管内皮细胞被受体内皮细胞替代(内皮嵌合)的现象,并分析内皮嵌合对肾脏移植物预后的影响。方法:移植后2周到4个月男性供者、女性受者的移植肾穿刺活检标本34例,选择Y染色体长臂Yq12区域(异染色质区)DNA片段作为探针,同时选择X染色体着丝粒区(α卫星DNA)探针作为对照,通过在石蜡切片标本上进行间期细胞双色荧光原位杂交,分析血管内皮细胞被受体替代的现象,分析其对移植肾预后的影响。结果:肾脏移植物中血管内皮细胞嵌合现象较普遍存在,内皮细胞的分布呈灶状,供者内皮细胞和受体内皮细胞可相邻存在。血管内皮细胞嵌合的发生与否移植肾1年血清肌酐值分别为(115.88±25.06)μmol/L,(118.16±18.85)μmol/L,差异无显著性意义(P>0.05)。结论:肾脏移植物中血管内皮细胞可以被受者来源的内皮细胞所替代,对移植后1年的肾功能无明显影响,内皮嵌合对移植肾的长期存活的影响有待进一步观察。     

慢性移植肾肾病管周毛细血管内皮细胞嵌合现象分析

目的:观察慢性移植肾肾病(CAN)中血管内皮细胞被受者内皮细胞替代(内皮嵌合)的现象,分析内皮嵌合对CAN进展的影响。方法:无功能移植肾切除组织,符合慢性移植肾肾病(CAN)造成移植肾失功、男性供者女性受者标本29例,选择Y染色体长臂Yq12区域(异染色质区)DNA片段作为探针,同时选择X染色体着丝粒区(α卫星DNA)探针作为对照,通过在石蜡切片标本上进行间期细胞双色荧光原位杂交(FISH),观察肾小管周毛细血管(PTC)内皮细胞的嵌合程度。结果:CAN中PTC内皮细胞嵌合现象较普遍存在,内皮细胞的分布呈灶状,供者内皮细胞和受体内皮细胞可相邻存在。PTC密度与间质纤维化程度有关,CD31阳性的PTC每单位面积密度14.52±4.85个,微血管内皮细胞嵌合出现比例为0.16±0.04,两者无明显相关性(r=0.13,P>0.05)。结论:肾脏移植物中血管内皮细胞可以被受者来源的内皮细胞所替代,CAN中PTC密度减少,内皮细胞的嵌合的发生与CAN的进展无直接关系。     

Preimplantation gender diagnosis by fluorescence in situ hybridization

目的：应用荧光原位杂交技术进行人类胚胎植入前性别诊断。方法：对2例甲型血友病基因携带、1例男性葡萄糖6磷酸脱氢酶（G-6-PD）缺乏患和2例Y染色体异常的患进行了6个周期的超排卵治疗。胚胎活检后取单个细胞进行固定,然后用荧光原位杂交技术检测胚胎的性别,根据遗传学规律选择胚胎性别后将胚胎移植入子宫。结果：5例患经6个治疗周期共取卵123个。可供活检的胚胎共61个,活检成功率为86.9％,活检后继续分裂率为62.3％,活检细胞固定率为98.1％。我们共移植了16个女性胚胎和3个男性胚胎,获得1例生化妊娠和3例临床妊娠,分别在羊水细胞和减胎组织中证实诊断的准确性。结论：荧光原位杂交技术用于遗传病的种植前诊断准确有效。对血友病等进行植入前性别诊断能避免选择性流产和重型患儿的出生,有利于优生。     

Cytogenetic characteristics of B cell chronic lymphocytic leukemia in 275 Chinese patients by fluorescence in situ hybridization: a multicenter study

Background  Under conventional cytogenetic (CC) analysis, only 30%–50% of B cell chronic lymphocytic leukemia (B-CLL) cases show clonal aberrations. Using fluorescence in situ hybridization (FISH), the percentage of patients with abnormalities rises to almost 80%, among them, the most frequent abnormalities were 13q14, 11q22, p53 deletions and trisomy 12. The aim of this study was to explore the incidence of cytogenetic changes in Chinese patients with B-CLL.

Methods  We used FISH methods to detect the cytogenetic features in 275 cases of B-CLL from 48 hospitals. The correlation between FISH abnormalities and clinical characteristics such as age, gender, white blood cell count, peripheral hemoglobin (Hb) level, peripheral platelet count (PLT), lactate dehydrogenase (LDH) level, Rai stage, Binet stage, and overall survival was analyzed, and the relationship between them and overall survival was also analyzed to evaluate their prognostic implications.

Results  Of the 275 patients, genetic aberrations were found in 77.8% using FISH. The frequencies of abnormalities were as follows: 13q deletion (56.4%), trisomy 12 (34.5%), p53 deletion (33.5%) and 11q22 deletion (30.5%). It was obvious that the patients with p53 deletion had lower level of Hb (P=0.001) and PLT (P=0.003) when compared to patients without p53 deletion. Significant differences were obtained in the distribution of p53 deletion according to Rai and Binet classification systems (P=0.016 and 0.008 respectively). Significant differences were also observed when the overall survival was correlated with p53 deletion (P=0.043), Rai stage (P=0.006), Binet stage (P=0.013), Hb level (P=0.004) and PLT level (P=0.010).

Conclusions  Chinese CLL patients have the similar frequencies of del(13q), trisomy 12, del(11q) and a higher frequency of del(17p) when compared to literatures. Del(17p) is associated with advanced stage and low levels of Hb and PLT. Patients with p53 deletion, or advanced stage probably have poor survival in China.

     

用荧光原位杂交技术显示染色体易位

目的：应用荧光原位杂交技术（ｆｌｕｏｒｅｓｃｅｎｃｅ ｉｎ ｓｉｔｕ ｈｙｂｒｉｄｉｚａｔｉｏｎ,ＦＩＳＨ）对Ｇ显带提示有染色体易位的病例进行分析,阐明易位本质。方法：以定位于待研究染色体区段的酵母人工染色体（ｙｅａｓｔ ａｒｔｉｆｉｃｉａｌｃｈｒｏｍｏｓｏｍｅ,ＹＡＣ）作为ＤＮＡ来源,结果：两组经Ｇ显带未能明确显示的染色体结构异常,经ＦＩＳＨ证实,一例为发生于１１号染色体与１３号染色体之间的平衡     

荧光原位杂交技术在膀胱肿瘤诊断中的应用进展

早期诊断膀胱肿瘤,提高患者的预后一直是泌尿外科医师思考的问题。近年来分子细胞遗传学快速发展,荧光原位杂交（Fluorescence in situ Hybridization Technique,FISH）技术已广泛应用于临床和科研检验。其在膀胱肿瘤诊断中以快速、无创、高特异性和高敏感性而引起广大研究者极大的兴趣。现在的研究证明,膀胱癌复发与染色体畸变及相应的基因变化有关,是由确定的分子事件促发的。本文对这方面的研究进行综述。     

应用荧光原位杂交分析移植山羊体内的人源细胞

目的 应用荧光原位杂交(flurescence in situ hybridization,FISH)分析人造血干细胞(human hematopoietic stem cell,hHSC)经宫内移植至山羊体内后人源细胞的存在、比例及其动态变化。方法 用人特异17号染色体卫星DNA探针对13头移植山羊进行间期FISH(interphase FISH,ISISH)分析,并用人男性特异Y染色体卫星DNA探针对其中2头移植人男性HSC的山羊进行IFISH分析。标本采用外周血细胞滴片、骨髓涂片和肝脏组织印片。结果 在13头移植山羊中,11头山羊体内有人源细胞存在,其中2头为人男性细胞,证实这些山羊为人／山羊异种移植嵌合体;人源细胞比例随山羊月龄递减,最长已存活21个月;所检测月龄段的移植山羊外周血、骨髓和肝脏组织中人源细胞比例小于1‰,但其中2头山羊肝脏组织中人源细胞集中一处,局部比例高达207.92‰和392．41‰。结论 FISH检测出人源细胞能在山羊体内存在并长期存活,表明山羊是经宫内移植hHSC的合适受体;人源细胞在外周血和骨髓中比例低,而在肝脏组织局部比例高,提示山羊肝脏组织局部内环境适于人源细胞存活、增殖和分化。     

用荧光原位杂交技术进行染色体异常者胚胎植入前遗传学诊断

目的：初步探索应用荧光原位杂交(FISH)技术为染色体异常患者进行胚胎植入前遗传诊断(PGD)。方法：针对染色体结构异常的种类．分别选择亚端粒探针及着丝粒探针，用荧光原位杂交技术为5例染色体异常患者进行植入前遗传学诊断。结果：5例患者共行7个PGD周期，共获卵134个，活检47个胚胎，FISH分析可供移植胚胎16个，6个移植周期移植其中15个胚胎，获得1例临床妊娠，并经产前诊断验证，已顺利诞生1健康男婴。结论：荧光原位杂交技术能有效地应用于染色体异常者胚胎植入前遗传学诊断。     

Chromosomal changes detected by fluorescence in situ hybridization in patients with acute lymphoblastic leukemia

Objectives To investigate patients with acute lymphoblastic leukemia (ALL) for TEL/AML1 fusion,BCR/ABL fusion, MLL gene rearrangements, and numerical changes of chromosomes 4, 10, 17 and 21 by fluorescence in situ hybridization (FISH) and to determine the relationship and the significance of those findings.Methods Fifty-one American patients (34 men and 17 women) were included in this study. Of them there were 41 patients with pro-B cell type ALL, 9 with B cell type ALL and 1 with T cell type ALL.Chromosome metaphases of each sample were prepared according to standard protocols.Fluorescence in situ hybridization was performed using commercially available DNA probes, including whole chromosome painting probes, locus specific probes, specific chromosome centromere probes and dual color/multiple color translocation fusion probes. The digital image analysis was carried out using Cytovision and Quips FISH programs.Results An overall incidence of chromosomal anomalies, including t (9; 22 ), MLL gene rearrangements, t (12;21), and numerical chromosomal anomalies of chromosomes 4, 10, 17 and 21 was found in 33 patients (65%). Thirty-one of them were pediatric patients and two adults. The t(12;21) was the commonest chromosomal anomaly detected in this population; 14 out of the 45pediatric patients (31%) were positive for TEL/AML1 fusion, among which three had an additionalderivative 21[t (12;21) ], four had a deletion of 12p and two had an extra copy of chromosome 21.All 14 patients with positive TEL/AML1 fusion had ALL pre-B cell or B-cell lineage according to standard immunotyping. The percentage of cells with fusion signals ranged from 20% to 80%. All fourteen patients positive for TEL/AML1 gene fusion were mosaic. Three out of the 14 patients positive for the TEL/AML1 gene fusion were originally reported to be culture failures and none of the remaining eleven samples had been found to have chromosome 12 abnormalities by conventional cytogenetic techniques. All pediatric patients with pre-T or T cell lineage and the six adults were negative for TEL/AML1 fusion. One patient had double Philadelphia chromosomes, three had a rearranaement or a deletion of the MLL aene. one had t (4;11)and two had a deletion of the MLL One of the patients with an MLL deletion also had a large ring of chromosome 21, and r (21) was caused by AML1 gene tandemly duplicated at least five times. The second case with the MLL deletion was also unique, the patient had a t (12;21) as well. A total of 20 patients had numerical changes( gain or loss) of chromosomes 4, 10, 17 and 21. Eight patients were found to have trisomies of three or four different chromosomes. Interestingly, seven of these patients did not have TEL/AML1, BCR/ABL or the MLL aene rearranaement, one did have the TEL/AML1 aene fusion. Eleven patients with pro-B cell or B cell type ALL (9 children with ALL, 2 adults with ALL) had numerical changes of chromosome 21 (gain 1 or 2 chromosome 21 ), among them, 10 patients had no structural alteration of chromosome 21, and one was combined by t (12;21 ). Four patients had a monosomy of chromosome 17 and three out of these patients with monosomy 17 also had a fusion signal of TEL/AML1.     

应用荧光原位杂交技术检测肺癌针吸标本中超二倍体肿瘤细胞

目的：探讨肺癌针吸标本中肿瘤细胞染色体数目的异常变化。方法：用７号、１１号、１７号、Ｘ梁色体特异的着丝粒ＤＮＡ探针,对３０例肺癌针吸标本（６例为肺癌原发灶标本,２４例为颈淋巴结标本）进行双色荧光原位杂交。结果：２７例肺癌阳性针吸标本中７号、Ｘ、１７号、１１号染色体超二倍体的病例分别达到８１．５％（２２／２７）、７７．８％（２１／２７）、７０．４％（１９／２７）、６３．０％（１７／２７）。３例肺癌阴性针吸标本中４个染色体均未出现超二倍体改变。结论：⑴用荧光原位杂交技术对针吸标本进行细胞遗传学分析是一种可行、简单的方法。⑵肺癌针吸标本中肿瘤细胞染色体超二倍体改变对诊断良、恶性细胞具有一定的参考价值。     

荧光原位杂交技术在膀胱癌诊断中的应用价值

目的 探讨荧光原位杂交技术(FISH)在膀胱癌诊断中的价值,并与常规的尿脱落细胞学对比.方法 收集100例疑似尿路移行上皮细胞肿瘤的血尿患者在行膀胱镜检查之前的尿液标本,分别做尿细胞学检测和荧光原位杂交分析.20例正常人尿液标本,用于建立FISH阈值,作为阳性判断的标准.结果 细胞学和FISH的总体灵敏度分别为43.2%、82.4%,特异度分别为:92.3%、88.5%,诊断符合率分别为:56.0%、84.0%,细胞学和FISH在灵敏度、诊断符合率、非浸润性肿瘤和初发肿瘤等方面的差异均有统计学意义.结论 在膀胱癌诊断中FISH较常规细胞学具有较高的灵敏度和诊断符合率,更易检出早期肿瘤.     

荧光原位杂交技术的应用

荧光原位杂交（fluorescence in situ hybridization, FISH））技术是一门新兴的分子细胞遗传学技术,在疾病诊断中发挥了重要作用。本文主要对FISH技术的原理特点以及临床应用作一综述。     

小鼠Connexin 31基因组克隆的筛选及荧光原位杂交定位

Connexin 31 (Cx31 )是人类遗传性疾病基因 ,利用小鼠Cx31基因的cDNA序列探针从小鼠的基因组文库筛选出Cx31的gDNA克隆 ,并用荧光原位杂交技术将其定位于小鼠 4号染色体的中部 ,为小鼠Cx31的基因结构和功能的研究以及转基因小鼠和基因剔除小鼠的研究打下了良好的基础     

荧光原位杂交在染色体异常产前诊断中的应用研究

目的对国产荧光原位杂交（FISH）试剂盒在染色体异常产前诊断中应用的可行性进行评估。方法对115例羊水样本行常规染色体核型检测外,进行间期FISH快速检测。采用国产FISH探针,特异性识别21、18、13、X、Y染色体。结果 115例样本中FISH杂交成功100例,结果获取需24～48h,其中95例为正常;3例为21三体;1例为18三体,结果与染色体核型分析一致;另有1例异常细胞数比例未达到判定阈值而结果不能判定,该样本的核型分析显示为47,XXY[7]/46,XY[43]的低比例嵌合体。未见假阴性和假阳性。结论国产FISH试剂盒具有较好的敏感性和特异性,与核型分析相比具有用材少、耗时短、操作简便及人员专业性依赖程度小的优势,但存在探针靶点有限、实验操作失败和结果不能判定的概率较高等问题。     

荧光原位杂交在染色体异常产前诊断中的应用研究

目的 对国产荧光原位杂交(FISH)试剂盒在染色体异常产前诊断中应用的可行性进行评估.方法 对115例羊水样本行常规染色体核型检测外,进行间期FISH快速检测.采用国产FISH探针,特异性识别21、18、13、X、Y染色体.结果 115例样本中FISH杂交成功100例,结果获取需24～48 h,其中95例为正常;3例为21三体;1例为18三体,结果与染色体核型分析一致;另有1例异常细胞数比例未达到判定阈值而结果不能判定,该样本的核型分析显示为 47,XXY[7]/46,XY[43]的低比例嵌合体.未见假阴性和假阳性.结论 国产FISH试剂盒具有较好的敏感性和特异性,与核型分析相比具有用材少、耗时短、操作简便及人员专业性依赖程度小的优势,但存在探针靶点有限、实验操作失败和结果不能判定的概率较高等问题.     

荧光原位杂交检测龈下菌斑中未获培养微生物

目的在龈下菌斑中观察一种未获培养的牙周可疑致病微生物AU126。方法针对AU126设计并评估特异性探针。使用激光共聚焦显微镜,通过与包含插入子克隆的原位杂交,定量分析荧光信号强度研判杂交严谨度。在严谨度分析获得的特定条件下,将慢性牙周炎患者的龈下菌斑标本与AU126探针进行杂交,运用共聚焦显微镜寻找AU126。结果AU126探针理想杂交是在10%甲酰胺中进行。通过对细菌轮廓荧光强度均值的判断,证实龈下菌斑中存在ROX-126标记的阳性细菌。细胞呈杆状,长4.0~5.4μm,宽0.64~0.84μm。结论在评估了AU126探针的特异性以及确定理想杂交严谨度的条件下,运用荧光原位杂交结合激光共聚焦显微镜技术,成功观察到人类龈下菌斑中的一种未获培养微生物AU126。     

荧光原位杂交技术检测对复发性膀胱癌的诊断价值

目的:探讨荧光原位杂交(FISH)技术在膀胱癌复发检测中的价值。方法:经尿道膀胱肿瘤电切或者钬激光切除术后患者49例,分别采用膀胱镜、FISH技术和尿脱落细胞学方法检测膀胱癌复发情况。结果:检测结果发现,膀胱癌术后复发31例。FISH技术、尿脱落细胞学检查对膀胱癌复发诊断的特异性均为100%。FISH技术检测敏感性为83.9%,高于尿脱落细胞学检测的敏感性(38.7%),差异有统计学意义(P<0.05)。结论:FISH技术较细胞学检测对诊断膀胱癌复发更为有效,具有广阔的临床应用前景。     

荧光原位杂交在产前诊断中的应用

目的 分析荧光原位杂交分析技术（fluorescence in situ hybridization,FISH）在快速诊断羊水细胞染色体异常中的应用价值。方法 以2013年6月至2017年12月间在首都医科大学附属北京妇产医院产前诊断中心接受羊水穿刺的1 676名孕妇为研究对象,抽取羊水后同时进行荧光原位杂交分析和染色体核型分析,然后比对两种检测的检测结果。结果 共收集1 676名同时进行核型分析和FISH检测的孕妇检测结果。羊水穿刺指征包括：高龄（637例,38.1%）、唐氏筛查高危（699例,41.71%）、无创游离DNA检测高风险（315例,18.79%）、B超异常（16例,0.95%）等。FISH检测结果阳性240例（14.13%）,包括21三体140例、18三体44例、13三体14例、性染色异常32例以及嵌合体10例。核型分析阳性252例,除包括上述240例FISH检测阳性结果外,还包括22号染色嵌合体1例、平衡结构重排6例、非平衡结构重排1例以及多态4例。结论 FISH检测能够快速准确诊断21号、18号、13号以及性染色体数目异常,根据FISH结果可进行进一步临床处理。怀疑胎儿携带其他染色体异常时,需联合核型分析等方法综合使用。     

用荧光原位杂交技术快速诊断孕早期胚胎染色体数目异常

目的探讨荧光原位杂交(FISH)技术在快速产前诊断孕早期胚胎染色体数目异常中的价值.方法采用13,18,21,X和Y染色体特异性DNA探针,对46例孕47～65 d高危孕妇的绒毛间期细胞进行FISH检测,同时行常规染色体核型分析平行诊断.结果与染色体核型分析结果一致的染色体数目正常43例,异常3例(47,XY 21;47,XY 18和45,X).3例异常核型胚胎经治疗性流产后再分别对其绒毛行常规染色体核型分析,结果与产前诊断相符.结论FISH技术用于产前诊断孕早期胚胎染色体数目异常具有快速、准确等优点.     

荧光原位杂交技术在检测人精子染色体异常中的应用进展

荧光原位杂交技术(FISH)以其快速、简便、高灵敏度及高特异性的优点,可一次检测成千上万精子染色体而被广泛应用,并逐渐成为检测精子染色体的首选方法.本文总结近10年来国内外相关文献,将FISH技术在检测正常人、不育者、染色体平衡易位者、有毒物接触者及肿瘤患者化疗后精子染色体异常方面的应用加以综述.