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目的:建立泻痢康肠溶胶囊的微生物限度检查方法.方法:依据《中国药典》2015年版四部通则1105、1106及1107和《中国药品检验标准操作规范》(2010版)收载的微生物限度检查方法对该药品微生物限度检查方法进行适用性试验.结果:需氧菌总数检查时,需要把供试品稀释到10-3;霉菌和酵母菌总数检查时,需要把供试品稀释到10-2;大肠埃希菌常规法即可;耐胆盐革兰阴性菌检查时需要增加培养基体积至15 ml;沙门菌检查时需要增加培养基体积至200 ml.结论:需氧菌总数和霉菌和酵母菌总数计数法采用的是供试品溶液稀释法;大肠埃希菌采用的是常规法;耐胆盐革兰阴性菌和沙门菌采用的是增加培养基体积的方法,采用上述方法,回收比值均在0.5~2范围内. 相似文献
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目的 应用siRNA下调胶质母细胞瘤TJ-905细胞株Notch-1基因抑制肿瘤增殖活性.方法 实验设以Notch-1为基因靶点的siRNA1、siRNA2、siRNA3转染组、阴性对照组和空白对照组,采用OligofectamineTM2000二次转染方法将 siRNA转移到细胞内.实时荧光定量PCR检测siRNA对Notch-1基因表达的抑制作用,筛选出最优siRNA序列用于后续研究;四唑盐(MTT)比色法检测细胞增殖活性;使用Notch-1 siRNA或阴性siRNA转染的TJ-905细胞悬液注射裸鼠腋下,观察移植瘤生长情况,比较肿瘤体积,观察3周后处死动物,切片行HE染色鉴定.结果 (Ⅰ)siRNA明显抑制Notch-1 mRNA的表达水平,空白对照组、阴性对照组、siRNA1、2、3转染组Notch-1 mRNA 表达率分别为1.00±0.07、1.04±0.05、0.11±0.02、0.12±0.01、0.77±0.03,siRNA1为最优序列.(2)siRNA1转染组细胞增殖活性显著降低.(3)接种Notch-1 siRNA1转染的TJ-905细胞裸鼠肿瘤生长速度明显低于接种阴性转染的TJ-905细胞裸鼠,裸鼠处死后观察肿瘤体积:Notch-1 siRNA1转染组为(748.3±154.3)mm3,阴性转染组为(1739.2±249.7)mm3,病理切片HE染色证明为多形性胶质母细胞瘤.结论 化学合成的靶向Notch-1基因的siRNA可以显著地下调Notch-1基因表达水平,抑制肿瘤增殖活性.为进一步进行胶质瘤的基因治疗研究奠定基础.Abstract: Objective To study the inhibitory effect of siRNA on glioblastoma (GBM) Notch-1 gene expression in addition to the growth of TJ- 905 glioblastoma.Methods Three Small interference RNAs (siRNAs) targeting Notch- 1 gene named siRNA1,siRNA2,siRNA3 were synthesized chemically in vitro with gene bank BLAST.TJ-905 cells were transfectedtwice with the siRNA by using OligofectamineTM2000.The nontransfected cells and nonspecific siRNAs transfected cells were taken as blank and nonsense controls.Down - regulation of Notch- 1 was demonstrated by real- time RT- PCR,according to the result of QRT- PCR we therefore selected the most effective siRNA for further study.Cell proliferation was measured by MTT analysis.Male Balb/C nude mice were injected subcutaneously with Notch- 1 siRNA- or nonsense siRNA- transfected TJ-905 cells,tumor sizes were measured every 4 days.HE stain was used to determine the property of tumor.Resuts(1) The expression of Notch- 1 mRNA in blank,nonsense controls,siRNA1,2,3 groups were 1.00±0.07,1.04±0.05,0.11 ±0.02,0.12 ±0.01,0.77 ±0.03 by real-time RT- PCR scan analysis,the most effective siRNA was siRNA1.(2) An inhibitory proliferation and growth can be induced in siRNA1 transfected group.(3)Nude mice xenografted with cell suspensions from the nonsense siRNA group developed tumors with a significantly increased volume (from the 18th days) as compared to mice that received the Notch- 1 siRNA1- treated cells.The final tumor volume were less in nude mice injected with Notch- 1 siRNA (748.3 ±154.3 )mm3 compared to nonsense si RNA injection (1739.2 ± 249.7 )mm3,HE stain demonstrate that tumor was multiformity glioblastoma.Conclusion The chemically synthesized siRNA targeted Notch- 1 gene could down -regulate the expression of Notch- 1.In addition,an inhibitory proliferation and growth were induced when compared with the control cells in vitro and in vivo.It was suggested that the suppression of Notch- 1 expression and the inhibition of growth provide a new way to glioma gene therapy. 相似文献
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靶向Notch-1基因的siRNA对人脑胶质母细胞瘤细胞裸鼠体内成瘤性的影响 总被引:1,自引:0,他引:1
背景与目的:近些年的研究发现,Notch-1基因在人脑胶质母细胞瘤的发生、发展中起到了重要的作用.本研究旨在观察靶向Notch-1基因的小干扰RNA(siRNA)对人脑胶质母细胞瘤裸鼠体内成瘤性的影响.方法:以OligofectamineTM为载体将靶向Notch-1基因的SiRNA转染人脑胶质母细胞瘤细胞系TJ905中,并设立无意序列siRNA(nonsense siRNA)转染组.然后,把经Notch-1 siRNA和经无意序列siRNA转染的细胞及未经转染的细胞分为3组.每组细胞悬液注射裸鼠5只,注射量按1×107/mL无血清培养基注射入每只裸鼠皮下200 μL,观察动物的成瘤情况,每隔3 d记录-下裸鼠移植瘤的体积V=(ab2)/2(a为肿瘤的长径,b为肿瘤的宽径),并在接种成瘤后对各组动物进行相应的SiRNA/Oligofectamine TM混合物的瘤内注射治疗,对照组注射等量的PBS.观察20 d后处死动物,取出肿瘤组织,常规石蜡包埋,组织切片HE染色并进行Notch-1的免疫组化染色测定.结果:裸鼠处死后观察肿瘤直径:Notch-1 siRNA转染组为(1 203±206)mm3,siRNA转染组为(2 241±401)mm3,对照组为(2 309±466)mm3,Notch-1 siRNA转染组与无意siRNA转染组,以及Notch-1siRNA转染组与对照组之间比较均有统计学意义(P<0.05).免疫组化结果显示:Notch-1的表达在Notch-1siRNA转染组与其他两组之间的比较也同样具有统计学意义(P<0.01).结论:靶向Notch-1基因的siRNA可以通过有效地降低肿瘤细胞Notch-1的表达来抑制裸鼠皮下胶质母细胞瘤移植瘤的生长,但还需要对siRNA的合理使用剂量与转染方式进行更加深入的研究. 相似文献
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目的:建立虎掌南星的微生物限度检查方法,考察该饮片微生物污染的程度,并评估其潜在的风险。方法:依据《中国药典》2020年版四部通则1105、1106、1108收载中药饮片微生物限度检查法对虎掌南星进行方法适用性试验,对20个批次药品的需氧菌总数(TAMC)、霉菌和酵母菌总数(TYMC)、耐热菌总数(NAIRE)、耐胆盐革兰阴性菌、大肠埃希菌、沙门菌、金黄色葡萄球菌、铜绿假单胞菌、白念珠菌进行检查,同时对耐热菌做探索性检验。结果:需氧菌总数在102~107之间;霉菌和酵母菌总数在101~105之间;12批次检出耐热菌,检出的耐热菌数在101~103之间;15批次检出耐胆盐革兰阴性菌,N均小于104;13批次检出大肠埃希菌;0批次检出沙门菌;20批次均检出金黄色葡萄球菌;7批次检出铜绿假单胞菌;6批次检出白念珠菌;耐热菌数随水浴时间增加,菌数下降明显。结论:通过对微生物的污染调查分析,给虎掌南星饮片的合理使用提供了一些数据上的保障。建议根... 相似文献
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