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1.
目的 建立以纳米金颗粒为载体的DNA探针杂交方法,快速、特异地检测耐甲氧西林金黄色葡萄球菌(MRSA)特异性mecA基因.方法 采用直径为60nm的胶体金纳米颗粒与巯基修饰的2条mecA基因寡核苷酸探针共价结合,制备成固化的DNA 金纳米探针.将金纳米探针与其互补的mecA基因靶序列在液相中杂交,通过调节2条探针的比例和不同的检测方法对杂交反应条件和产物检测进行优化.结果 金纳米探针呈稳定的酒红色,与互补的靶序列DNA进行液相杂交后颜色呈现明显的蓝色变化.当2条探针不对称加入时颜色变化更明显.采用反向色谱反应盘检测反应产物也可获得满意的效果.产物离心后检测灵敏度可达2.36 fmol/L.结论 固化纳米胶体金探针杂交技术具有快速简便的特点,有望为MRSA的快速检测提供一种新的方法.  相似文献   

2.
金纳米颗粒基因探针同步检测HBV和HCV的初步研究   总被引:1,自引:0,他引:1  
目的 建立一种简单、快速同时检测HBV DNA和HCV RNA的诊断方法.方法 利用PCR方法扩增HBsAg阳性合并抗HCV阳性的患者血清中HBV DNA和HCV cDNA并提取.制备Au纳米颗粒HBV DNA、HCV cDNA基因探针.在含有Au纳米颗粒HBV DNA、HCV cDNA基因探针的液相检测体系中加入HBV DNA和(或)HCV cDNA,透射电镜下观察.结果 PCR(可扩增HBV DNA和HCV cDNA.出现预期的431bp和323bp特异性条带.使用Au纳米颗粒基因探针通过透射电镜可同步检出乙、丙型肝炎合并感染患者血清中提取的HBV DNA和HCV RNA.结论 通过透射电镜,使用Au纳米颗粒基因探针可同步检出乙、丙型肝炎合并感染患者血清中的HBV DNA和HCV cDNA,甚至可达到无需PCR水平,此方法具有敏感性高、特异性好的特点.  相似文献   

3.
目的 制备HBV共价闭合环状DNA(cccDNA)功能化磁性纳米颗粒,为进一步建立富集分离和检测cccDNA方法奠定基础. 方法 通过亲和素修饰纳米颗粒表面,利用亲和素与生物素的亲和作用,将亲和素修饰的纳米颗粒与生物素标记的cccDNA特异性探针偶联,制备cccDNA功能化纳米颗粒,最后用cccDNA探针完全互补的荧光标记序列与功能化纳米颗粒反应,鉴定其特性. 结果 亲和素修饰的纳米颗粒能结合生物素最大量为1 205 ng/mg;通过与亲和素作用,生物素标记cccDNA特异性探针固定在磁性纳米微粒表面,其最大结合生物素标记探针量为3.2×10-9 mol/mg;cccDNA功能化纳米颗粒可捕获与其互补的荧光标记寡核苷酸序列,其最大捕获量为2.0×10-9mol/mg,并能通过变性释放出保持生物学活性游离的荧光标记寡核苷酸序列. 结论 成功构建的cccDNA功能化纳米微粒能有效捕获与其互补的序列,其捕获量能达到cccDNA的分离提取要求.  相似文献   

4.
纳米金标记法在DNA检测及基因芯片技术中的应用   总被引:6,自引:0,他引:6  
近年来DNA介导的纳米金颗粒的组装研究受到广过重视,烷疏基修饰的寡核苷酸可与纳米金共价结合,形成的探针可与目的多核苷酸序列杂交,纳米金颗粒被可逆组装成超分子结构,聚集过程伴有红色到蓝色的颜色变化,由此衍生出比色法检测特定DNA序列的新方法。该方法具有极高的选择性和较好的灵敏度,与银染法结合更提高了检测的灵敏度,可用于斑点亲交、芯片检列等领域。本文对该方法的原理、操作过程及研究进展作一简要介绍。  相似文献   

5.
吴蓉  府伟灵  陈鸣 《重庆医学》2004,33(8):1135-1137
目的探讨用PNA作为探针与普通的DNA探针相比对压电传感器基因杂交的优越性.方法用生物素-亲和素法分别将PNA、DNA两种探针固定在基因传感器的金膜表面,分别与完全匹配、1个碱基错配、2个碱基错配的互补靶序列进行杂交,观察2种不同的探针与靶序列反应所引起的频率变化及所用的时间.结果相对于DNA探针,PNA探针识别碱基错配的能力明显高,且杂交时间更短.结论 PNA与靶序列DNA结合有高度的特异性,选用PNA作探针,可提高压电基因传感器检测的特异性.  相似文献   

6.
目的:研制以金纳米微粒标记和银染色信号放大技术为基础的DNA芯片,用于快速检测表皮生长因子受体(EGFR)基因突变。方法:氨基修饰的寡核苷酸探针固定于醛基化玻璃片基制备DNA芯片,5′-生物素标记引物用于PCR扩增EGFR基因第18、19、20、21外显子片段,杂交后含互补序列的PCR产物结合于芯片,金纳米微粒借助表面包被的链亲和素识别标记于PCR产物5′-端的生物素而与之结合,再经银染色法放大杂交信号,依据芯片杂交结果判读EGFR基因突变,并用DNA直接测序法进行验证。结果:所制备的DNA芯片可快速检测肿瘤组织标本中EGFR基因突变,检测敏感性达到10-9 mol/L,可检出标本中5%的基因突变。结论:所制备的DNA芯片具有高特异性和敏感性,且操作简便,适用于临床肿瘤组织标本基因突变的快速检测。  相似文献   

7.
目的 探讨将HIV多抗与磁性纳米微粒偶联制备免疫磁性纳米微粒,用于HIV的靶向性加热治疗,并对其在交变磁场下的升温能力进行检测.方法 采用共沉淀法制备四氧化三铁纳米材料,选择氨基硅烷偶联剂对磁性纳米微粒进行修饰,通过一定浓度戊二醛活化将抗HIV多抗交联制备免疫磁性纳米微粒,并对其在交变磁场下的升温能力进行检测.结果 磁性纳米颗粒经氨基硅烷修饰后平均粒径为30 nm左右.戊二醛活化的磁性纳米微粒与抗HIV多抗具有较高的结合率,可在体外稳定保存,具有良好的交变磁场下升温能力.结论 抗HIV多抗免疫磁性纳米微粒的结合率、稳定性和交变磁场下升温能力可满足进一步的HIV靶向性加热治疗.  相似文献   

8.
目的研究纳米金及其标记细胞色素P450在811nm近红外荧光光谱的特性,并对其荧光增强机理进行深入的探讨。方法用不同量的柠檬酸三钠在煮沸搅拌条件下还原氯金酸制得不同粒径的纳米金,利用纳米金易与蛋白结合的性质标记细胞色素P450。用紫外-可见吸收光谱仪、投射电子显微镜以及荧光光谱仪,研究纳米金及其标记细胞色素P450的光谱性质。结果透射电子显微镜显示随着还原剂柠檬酸钠加入量的增加,形成的纳米金的颗粒逐渐减小。紫外-可见吸收光谱显示大颗粒纳米金只在250nm处有吸收,当纳米金粒径的减小至21nm时,在525nm处出现新的吸收峰。荧光光谱仪显示以538nm为激发波长,纳米金在811nm处有荧光发射峰,且标记蛋白后荧光强度增加。结论通过对纳米金近红外荧光性质的研究,可利用纳米金的近红外荧光性质作为探针检测细胞色素P450。  相似文献   

9.
双功能化胶体金纳米探针的制备及其特异性考察   总被引:1,自引:0,他引:1  
目的:制备一种新型的双功能化胶体金纳米探针,并考察其特异性.方法:选择二硫化物分子和甲型流感病毒(H1N1亚型)HA基因中特异性的23 mer寡核苷酸片段组装到胶体金颗粒表面,制成双功能化胶体金纳米探针(识别探针,颜色呈酒红色);利用磁性微球作为固相支持物,偶联另一段特异性H1N1寡核苷酸片段制成捕捉探针;两者与靶DNA(完全匹配DNA)结合,形成无色的捕捉探针-靶DNA-识别探针("三明治"夹心式复合物),并采用基质辅助激光解析离子化飞行时间质谱(MALDI TOF MS)测定.同时,在选择向体系中分别加入超纯水(作为空白对照)、完全匹配DNA、不完全匹配DNA及2条仅有1个碱基错配的DNA片段作为靶目标进行实验,观察体系颜色变化及质谱测定,考察探针的特异性.结果:捕捉探针和识别探针只能与完全匹配DNA结合,形成夹心式复合物,反应体系颜色由酒红色转变为无色,充分洗涤后加热去杂交,则由无色复呈酒红色,质谱检测出现信号(m/z 693);而其他靶目标(不完全匹配DNA及2条仅有1个碱基错配的DNA片段)和空白对照实验,不能与2种探针形成夹心式复合物,反应体系均没有颜色变化(仍呈现酒红色),充分洗涤后体系呈无色,加热去杂交体系仍呈无色,质谱检测未见信号(m/z 693).这说明胶体金纳米探针的特异性很高,可区分仅有1个碱基错配的靶目标.结论:本研究建立了一种新型的自组装双功能化胶体金纳米探针,其制备过程简单、特异性很高.此类型胶体金纳米探针在核酸分析,尤其是疾病早期诊断,寡核苷酸多态性分型等方面将有广泛的应用前景.  相似文献   

10.
目的 制备聚乙烯亚胺修饰纳米金基因载体并研究其理化性质的表征参数和体外转染效率.方法 通过化学还原法制备聚乙烯亚胺修饰的纳米金基因载体,用绿色荧光蛋白质粒(pAcGFP-N1)做报告基因,纳米基因载体可通过静电吸附的方式结合质粒DNA.用紫外分光光度计检测其吸收光谱,用透射电镜观察其形态特征,激光粒度分析仪测定其粒度分布、表面电位(Zeta电位),1%琼脂糖凝胶电泳检测该基因载体与质粒DNA的结合稳定性,CCK-8实验检测聚乙烯亚胺修饰纳米金基因载体及DNA-纳米金复合物对HEK293细胞的细胞毒性作用,通过荧光显微镜观察聚乙烯亚胺纳米基因载体介导pAcGFP-N1在体外培养的HEK293细胞中的表达,并分析其转染效率.结果 聚乙烯亚胺还原氯金酸可以得到带正电荷的纳米颗粒,呈单分散球形分布,其粒径为(12.3 ±3.3)nm.在pH =7.2时,Zeta电位为+(29.7±5.1)mV.1%琼脂糖凝胶电泳结果表明,当纳米金/质粒DNA≥0.5时,质粒DNA可完全结合到纳米金表面.体外转染实验表明,聚乙烯亚胺修饰纳米金基因载体能介导pAcGFP-N1转染HEK293细胞并在细胞中表达绿色荧光蛋白,其转染效率可达25%.结论 聚乙烯亚胺修饰纳米金是一种新型非病毒基因载体,具有转染效率高、对细胞毒性小等优势.  相似文献   

11.
Objective To observe blood pressure change with age in salt-sensitive teenagers whose salt sensitivity were determined by repeated testing.Methods Salt sensitivity was determined through intravenous infusion of normal saline combined with volume-depletion by oral diuretic furosemide in 55 teenagers. After five years, salt sensitivity was re-examined and subject blood pressure was followed up. Blood pressure changes in salt-sensitive teenagers were compared to that of non-salt sensitive teenagers over five years.Results After 5 years, the repetition rate of salt sensitivity determined by intravenous saline loading is 92.7%. In teenagers with salt sensitivity on the baseline, both the systolic blood pressure increments and increment rates were much higher than non-salt sensitive teenagers (12.7±12.1 mmHg vs. 2.8±5.2 mmHg, P< 0.01; 12.2%± 12.0% vs. 2.5% ±4.4%, P< 0.001,respectively). There was a similar trend for diastolic blood pressure (8.4 ± 6.4 mmHg vs. 3.7 ± 6.4 mmHg, P = 0.052; 13.2% ±10.6 % vs. 6.8%± 10.1%, P = 0.053, respectively).Conclusions Salt sensitivity determined by intravenous saline loading showed good reproducibility. Blood pressure increments with age were much higher in salt-sensitive teenagers than non-salt sensitive teenagers, especially in terms of systolic blood pressure.  相似文献   

12.
Increasing maternal age is the only etiological factor unequivocally linked to Down's syndrome in humans. The occurrence rate of newborns with Down's syndrome is about 1/220 in women over 35 years old. However, the occurrence rate in embryos fertilized in vitro, of the elder woman is unclear. Using FISH we screened the number of chromosome 21 in preimplanted embryos of 5 elderly women (average age, 38.4 years) to study the feasibility and necessity of screening trisomy 21 in embryos in patients over 35 years old at the in vitro fertilization (IVF) center.  相似文献   

13.
Real-time three-dimensional echocardiography (RT3DE)is a new ultrasound technique that enables dynamic threedimensional visualization and quantification of the heart in real time. Investigation of feasibility and methodology of RT3DE in determining left ventricular (LV) and right ventricular (RV) volumes, RT3DE was performed in 35 normal adults using Philips SONOS 7500 system with a 2-4 MHz matrix array transducer. The 60°×60° "pyramid" volume database was obtained and analyzed on a TomTec echo workstation. Both LV and RV volumes were calculated with four 3DE methods (i.e. apical 2, 4, 8, and 16-plane) through manually tracing ventricular endocardial borders in end diastole and end systole. Stroke volumes were then calculated. LV volume was also measured by 2DE Simpson's rule using GE VIVID 7 ultrasound machine.  相似文献   

14.
A clinical guideline for the therapeutic interventions of integrative medicine may be defined as a written document which states a series of recommendations on therapeutic interventions of integrative medicine for a special disease or condition. The guideline may provide assistance to medical professionals in making clinical decisions aimed at improving the clinical outcome of patients and reducing the costs of medical care(~'4~. Recommendations issued by a guideline should be based on the best available evidence in both Western and Chinese medicine. For fulfilling this purpose, the development of clinical guidelines for therapeutic interventions in the field of integrative medicine should follow scientific principles and undergo a rigorous processes.  相似文献   

15.
Objective:To probe into the influence of changes of ovarian hormones on the pathogenesis of the specific sub-type premenstrual syndrome(PMS)and reveal partial microcosmic mechanisms of adverse flow of liver-qi.Methods:Estradiol(E2)and progesterone(P)levels in serum were determined at different phases of menstrual cycle by radioimmunoassay.Results:In the group of PMS with adverse flow of liver-qi.the secretive peak value Of E2 and P at the follicular phase significantly decreased,and the secretive peak value at the luteal phase did not come into being.Conclusions:Low E2 and P secretive peak at the follicular phase and absence of secretive peak at the luteal phase is one of the microcosmic mechanisms of PMS with adverse flow of liver-qi.One of the pathophysiologic mechanisms of specific sub-type PMS is probably the continuous low level of E2and P.  相似文献   

16.
Journal of Nanjing Medical University (English Edition) JNMU, sponsored by Nanjing Medical University, was established in 1987. It is a bimonthly comprehensive English medical journal published locally and abroad.Since 2007, Journal of Nanjing Medical University (English Edition )was granted Elsevier the full publishing and distribution rights worldwide for the Electronic Edition, excluding the People's Republic of China.  相似文献   

17.
Objective: To evaluatel the value of D-dimers in patients with acute aortic dissection (AAD). Methods: This study consisted of 16 patients with AAD and 27 non-AAD patients. Serum D-dimets were measured by Sta-Liatest D-DI immunoturbidimetric assay. Results: D-dimer level was higher (P < 0.001) in patients with AAD(7.91 ± 5.52 μg/ml) than that in non- AAD group(1.57±1.24 μg/ml). D-dimer was positive (>0.4 μg/ml) in all patients with AAD and in 10 control group patients (37%). Among patients with acute AAD, D-dimers tended to be higher in Stanford A than in Stanford B (8.67 ± 4.31 μg/ml vs. 3.24±1.27 μg/ml, P <0.01). D-dimer values tended to be higher in more extended disease(3.84 ± 1.65 μg/ml, 8.57 ± 3.58 μg/ml and 11.87 ± 5.69 μg/ml in thoracic aorta, thoracic and abdominal aorta, thoracic and abdominal aorta and iliacal arteries, respectively, P < 0.05 for both 8.57 ± 3.58 and 11.87 ± 5.69 vs. 3.84 ± 1.65 ). Including the control group into the analysis, we found a sensitivity of 100%, a negative predictive value of 100%, and a specificity of 66% and a positive predictive value of 64% for D-dimer in diagnosis of AAD in our patients with suspected AAD. Conclusion: D-dimer was elevated in patients with AAD. A negative D-dimer test result could be useful in excluding AAD.  相似文献   

18.
Objective: To set up a simple and reliable rat model of combined liver-kidney transplantation. Methods: SD rats served as both donors and recipients. 4℃ sodium lactate Ringer's was infused from portal veins to donated livers,and from abdominal aorta to donated kidneys, respectively. Anastomosis of the portal vein and the inferior vena cava (IVC) inferior to the right kidney between the graft and the recipient was performed by a double cuff method, then the superior hepatic vena cava with suture. A patch of donated renal artery was anastomosed to the recipient abdominal aorta. The urethra and bile duct were reconstructed with a simple inside bracket. Results: Among 65 cases of combined liver-kidney transplantation, the success rate in the late 40 cases was 77.5%. The function of the grafted liver and kidney remained normal. Conclusion: This rat model of combined liver-kidney transplantation can be established in common laboratory conditions with high success rate and meet the needs of renal transplantation experiment.  相似文献   

19.
FOR anesthesiologis s ,treatingpostoperativepainhas alwaysbeen a problem.Althoughopioidshave been provedtobe effective,theirsideeffectscouldnotbeignored.With thedevelopmentofscienceand pharmacology,many drugs with aspectsof satisfactoryanalgesicefficacyand couldbe welltoleratedby patientshave been developed.And lornoxicamisone of them, which isa non-steroidalanti-inflammatorydrug (NSAID ), with analgesic, anti-infl-ammatory,andantipyreticproperties.Itseliminationhalf-time(3 to 5 hours) isle…  相似文献   

20.
Objective To measure Derpl and Blot5 allergen levels in asthmatics' homes in Hongkong.Methods Seventy houses were enrolled for a mite indoor environment study. Dust samples were obtained from two sites of each patients' house: bed and floor. Derpl and Blot5 levels were quantified by a two-site monoclonal antibody-based ELISA technique.Results The levels of Derpl allergens found in bed (geometric mean (GM) 3.43 μg/g of dust; 95%CI, 1.89-4.96 μg/g)and on the floor (GM 1.12 μg/g of dust; 95%CI, 0.71-1.53 μg/g) indicated significant differences (P=0.005). However, the levels of Blot5 allergens found in bed (GM 19.00 μg/g of dust; 95%CI, 0.89-38.90 μg/g) and on the floor (GM 6.14 μg/g of dust; 95%CI, 0.40-11.90 μg/g) showed no statistically significant difference. In addition, in regards to the exposure index for Derpl and Blot5 allergens found in bed and on the floor, 17.6% in bed and 8.6% on the floor had levels of Blot5 ≥ 10 μg/g of dust, higher than those obtained for Derp1 (7.2% and 0% in bed and on the floor respectively, P< 0.05); higher percentages in bed and on the floor (25.0% and 35.7%) were observed for levels of Blot5 =0 μg/g of dust as compared with Derpl in bed and on the floor (4.3% and 14.5% respectively, P< 0.05).Conclusions Derpl and Blot5 are the major allergens found in this regional study, Blot5 is a more potent allergen in Hongkong, probably reflecting the high level of exposure to Blomia tropicalis (Bt). Bt and Dermatophagoides pteronyssinus (Dp) allergens should be included for precise diagnosis and effective immuno-therapeutic treatment of mite allergy in Hongkong.  相似文献   

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