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The different fishes which are employed in the present studies are wild-type salmon, cultured salmon of freshwater and seawater, sea perch and fat greenling. The complete CT gene sequences of salmons are obtained by PCR amplification. The partial CT sequences of sea perch and fat greenling are obtained by in vitro cloning PCR method. Alignment of obtained CT sequences with other fish CT shows that CT appears to be well conserved among the same family. And the relative in taxonomy is far away, the similarity of different fish CTs is low. On the contrary, the closer the relative in taxonomy is, the higher the similarity of different fish CTs is. The sCT is expressed in pGEX-4T-X by recombinant form . We also succeed in the research of sCT expression alone by expression PCR. In addition, sCT antiserum is obtained using GST-sCT as antigen, and the high titer is tested by double immunodiffusion. In the rat bioassay, administration of 50 μg recombinant protein evoked significant hypocalcemia at 1 h after the data are analyzed by t-test.

本文用PCR方法克隆了野生鲑鱼、养殖鲑鱼的降钙素基因,并应用体外克隆PCR的方法首次克隆出鲈鱼、六线鱼降钙素的部分基因序列,通过对克隆的降钙素序列的比较研究,结果显示同一科的鱼降钙素序列保守性较高,同时,根据降钙素的部分氨基酸序列进行了降钙素相似性的比较研究,结果显示在分类学上,分类地位较远的鱼,其降钙素相似性较低,分类地位越接近的鱼,其降钙素相似性越高;我们利用谷胱甘肽S-转移酶(Glutathions S-transferase,GST)融合表达载体pGEX-4T-X对克隆的鲑鱼降钙素基因进行了融合表达研究,应用表达PCR的方法对降钙素基因的独立表达进行了初步的研究探索,并将纯化的融合蛋白作为抗原,获得了高效价的兔抗鲑鱼降钙素免疫血清;生物活性研究表明,大肠杆菌表达的融合蛋白具有显著的降血钙作用。

By retrovirus vector the foreign mdr1 gene could be efficiently transferred into bone marrow mononuclear cells of mouse in vitro, and it was confirmed that mdr1 gene was expressing stably and effectively in cells; in short terms there is no significant influence on the bone marrow MNC by transfection of mdr1 gene,but in the long run(in our study we had surveyed 2 months),there is a decline in the expression of apoptosis factor Bax in the MNC which transfected by mdr1 gene,in favour of the recovery of the receptor mouse;on the other hand it also poses a blance problem between multiplication and apoptosis in the process of receptor hematopoiesis after transplanting HSC which was transfected with mdr1 gene.

通过逆转录病毒载体可在体外将外源性mdr1基因转入小鼠骨髓单个核细胞中,转染的mdr1基因能整合到骨髓单个核细胞基因组中并有功能性表达;mdr1基因转入小鼠骨髓单个核细胞在移植后短期内对受体造血细胞没有显著影响,但从长期看来(本研究观察2个月)转染了mdr1基因的造血干细胞的凋亡因子Bax表达降低,更有利于受体鼠造血的恢复;另一个方面它也同时提出一个问题,即mdr1基因转染造血干细胞在回输受体后造血过程的增殖凋亡平衡。

In this study we observed the characters of cyclooxyenase-2 expressions and distributions in rat alveolar osteoblasts in vivo or in vitro in different force environments. And we tried to find the suitable mechanical environment to benefit the reproduction and remodification of alveolar bone in oral clinical treatment.

本研究目的在于通过观察体内、体外不同压力环境下牙槽骨成骨细胞 COX一2表达和分布的特点,探讨如何建立有利于牙槽骨新生和重建、减少牙槽骨吸收的合适力学环境,以期为口腔临床修复提供参考依据。

Effects of Adv-hBMP2 gene modify on the bone formation of tissue-engineered bone substitute. BMSCs of canine transferred by Adv-BMP2 gene expressed ALP in vitro and induced ectopic bone formation in nude mice. BMSCs adhered on the surface of FDB after 4hrs, expanded in 24hrs and proliferated in polylayer in 4d. FDB, combined with BMSCs transferred by Adv-BMP2 gene, induced bone formation in the subcutis of nude mice in 6 weeks.

结果:1.BMP-2基因修饰对组织工程化骨成骨的影响 BMSCs经Adv-BMP2基因转染后ALP染色强阳性、并在裸鼠肌肉形成异位骨组织;转基因的BMSCs与冻干骨复合后24小时即完全贴附生长,4天后复层生长并分泌大量胶原,植入裸鼠背部皮下6周后可见大量成骨,很少吸收,而单纯冻干骨或冻干骨复合BMSCs基本不成骨。2。

On the functional differentiation,we found and that the type II ras-GTPaseactivating protein (p100-GAP),which was specifically expressed in humanplacenta,was largely expressed in the highly-differentiated syncytiotrophoblastcells and moderately expressed in the cultured intermediate trophoblast cells.While in the non-differentiated cytotrophoblast cell line-NPC,no expression was observed.Its mRNA and protein expressingcharacterization was in consistent with that of hCGβ,which was one of the mostimportant markers of the trophoblast cell functional differentiation.In addition,the expressing amount of p100-GAP increased with the progressing of pregnancyand the syncytium formation in vitro.

细胞功能分化的研究结果表明:经重组质粒的构建获得特异表达于人胎盘的p100-GAP的cDNA探针后,从蛋白和分子水平证实p100-GAP大量表达于分化程度高的合体滋养层细胞,少量表达于体外培养的中间型滋养层细胞,而非分化的细胞滋养层细胞NPC则不表达,这种表达特性与作为滋养层细胞功能分化重要指标之一的hCGβ高度一致;p100-GAP表达量还随妊娠过程和体外合体化过程而逐渐增加。

DCs acquired by our reformed methods express both CD83 and CD 14 molecules highly, and have a higher density than other domestic reports. The higher TNF in DCs culture medium of HC patient suggests DCs in patient still have antigen presenting ability and by optimization the culture medium would improve its presenting ability and have a potential value in design and application individual vaccine. Although antigens pulsed DCs have a decrescent antigen presenting ability but BCG HSP70 could induce its mature and improve its presenting ability. Suggests BCG HSP70 would be a useful mature inducer. Lymphocytes primed by DCs based HC vaccine have the specific cytotoxicity against HCC lines. The CTL after freezing and anabiotic could prophylaxis and therapy HC xenograft on nude mouse. The results also suggests that CD4〓 lymphocytes play a important role in HC with a good differentiation and would be useful in treatment this kind of HC. After being activated by Peptide LLNQHACAV of hAFP and apoptotic HCCs pulsed DCs respectively, the culture medium of activated lymphocytes both contains a high level Th1 cytokines IL-12 and TNF. Primed lymphocytes appeared a characteristic of NK cells. DCs not only inhibited the growth of human HCC and other cancer cells in vitro but also prevented the growth of HC xenograft on nude mouse in vivo. There are at least three kinds of mechanism playing important role in DC based vaccine ,there are inhibition of DCs, HC specific CTL and cytokines pathway.

诱导出的DC共同表达CD83和CD14分子,CD83分子表达明显高于国内报道;肝癌患者DC培养上清中TNF水平高于健康人,提示肝癌患者DC仍具一定的抗原呈递能力,适当调控可使其行使APC功能,以期在肝癌个体化疫苗中发挥作用;DC负载肝癌可溶性抗原后,抗原呈递能力降低,BCG HSP70可促进DC成熟,增加其抗原呈递能力,预示BCG HSP70有可能成为促进DC活化和成熟的另一重要分子;肝癌DC疫苗在体外诱导肝癌特异性淋巴细胞,活化的淋巴细胞在体外对肝癌细胞的杀伤以特异性CTL为主,同时分泌较高水平Th1型细胞因子IL-12和TNF,并抑制4种肝癌细胞生长;冷冻复苏后的肝癌特异性淋巴细胞可以预防和抑制人肝癌裸鼠皮下移植瘤,提示DC负载肝癌可溶性抗原后诱发的MHC-Ⅱ类限制性CD4〓T细胞有可能在分化程度高的肝癌治疗中发挥作用;用DC和HLA-A2〓DC分别负载凋亡肝癌细胞和hAFP218-226位LLNQHACAV HLA-A2限制性九肽,在体外诱导肝癌特异性淋巴细胞,活化后的CTL细胞分泌较高水平的Th1型细胞因子IL-12和TNF,并具较强杀伤活性,此CTL同时具备NK细胞特征;DC对肿瘤细胞的抑制作用可能是通过吞噬实现的,Fas-L在DC抑制中也起一定作用;DC对人肝癌裸鼠皮下移植瘤的抑制率为97%;在肝癌DC疫苗的作用中,至少联合3种以上机制,即通过DC的直接作用、肝癌特异性CTL和细胞因子途径直接或间接地杀伤和抑制肝癌细胞。

The stable clones are further identified by RT-PCR and Western blot; 6 MTT assay is used to investigate the effect of ZNRD1 on the cell growth of cells (AGS, SGC7901, MKN28, NIH3T3, GES-1); 7 Soft agar assay is used to investigate the effect of ZNRD1 on the clonality of cells (AGS, MKN28); 8 Nude mice assay is used to investigate the effect of ZNRD1 on the cell growth of gastric cancer cells (AGS, MKN28); 9 Flow cytometry is used to investigate the effect of ZNRD1 on the cell cycle distribution of cells (AGS, MKN28, NIH3T3, GES-1); 10 Flow cytometry is used to investigate the effect of ZNRD1 on the cell apoptosis of cells (AGS, MKN28, NIH3T3); 11 MTT assay is used to investigate the effect of ZNRD1 on the drug sensitivity of cancer cells (SGC7901, SGC7901/VCR, HL-60, HL-60/VCR) in vitro; 12 SRCA is used to investigate the effect of ZNRD1 on the drug sensitivity of gastric cancer cells (SGC7901, SGC7901/VCR) in vivo; 13 Flow cytometry is used to investigate the effect of ZNRD1 on adriamycin accumulation of cancer cells (SGC7901, SGC7901/VCR, HL-60, HL-60/VCR); 14 Transmission electron microscope is used to investigate the effect of ZNRD1 on the sensitivity of SGC7901 cells towards drug-induced apoptosis; 15 Flow cytometry and DNA ladder assay are used to investigate the effect of ZNRD1 on the sensitivity of cells (SGC7901, SGC7901/VCR, HL-60/VCR) towards drug-induced apoptosis; 16 Microarray is used to investigate the profiling of ZNRD1-responsive genes in gastric cancer cells (AGS, MKN28, SGC7901, SGC7901/VCR); 17 RT-PCR and Western blot are used to identify the results of microarray; 18 Reporter gene assay is used to investigate the effect of ZNRD1 on the transcriptional activity of cyclin D1; 19 Reporter gene assay is used to investigate the effect of ZNRD1 on the transcriptional activity of MDR1; 20 Kinase assay is used to investigate the effect of ZNRD1 on the activity of cyclin E-CDK2 kinase; 21 The antisensenucleic acids of p21 is used to inhibit the expression of p21, and flow cytometry is used to investigate the effect of p21 on ZNRD1-induced cell cycle arrest in gastric cancer cells; 22 The antisensenucleic acids of p27 is used to inhibit the expression of p27, and flow cytometry is used to investigate the effect of p27 on ZNRD1-induced cell cycle arrest in gastric cancer cells; 23 Liposome is used to up-regulate the expression of Skp2, and flow cytometry is used to investigate the effect of Skp2 on ZNRD1-induced cell cycle arrest in gastric cancer cells; 24 Western blot is used to investigate the effect of ZNRD1 on the stability of Skp2 and p27 in gastric cancer cells; 25 MVD assay is used to investigate the effect of ZNRD1 on the angiopoietic activity of gastric cancer cells; 26 ELISA is used to investigate the effect of ZNRD1 on the expression of VEGF165 in gastric cancer cells; 27 The roles of DARPP-32 in MDR of gastric cancer cells are investigated using gene transfection, MTT assay, SRCA, flow cytometry and DNA ladder assay.

应用杂交瘤技术制备ZNRD1的首个单克隆抗体;2)利用RT-PCR、Western blot和免疫组化检测ZNRD1在胃癌组织、胃炎组织、正常胃上皮组织、胃癌细胞和正常胃组织上皮细胞中的表达;3)构建ZNRD1的小干扰RNA载体,并测序鉴定;4)利用脂质体将ZNRD1的真核表达载体及其空载体转染胃癌细胞(AGS、SGC7901、MKN28)和小鼠成纤维细胞(NIH3T3),G418筛选后进行鉴定;5)利用脂质体将ZNRD1的小干扰RNA载体及其空载体转染药敏胃癌细胞(SGC7901)、正常胃组织上皮细胞(GES-1)、对长春新碱耐药的胃癌细胞(SGC7901/VCR)、药敏白血病细胞(HL-60)、对长春新碱耐药的白血病细胞(HL-60/VCR),G418筛选后进行鉴定;6)利用MTT实验检测ZNRD1高/低表达对细胞(AGS、SGC7901、MKN28、NIH3T3、GES-1)生长的影响;7)通过软琼脂克隆形成实验检测上调ZNRD1对AGS、MKN28细胞克隆形成能力的影响;8)通过裸鼠成瘤实验检测上调ZNRD1对AGS、MKN28细胞体内成瘤性的影响;9)通过流式细胞仪分析ZNRD1高/低表达对细胞(AGS、MKN28、NIH3T3、GES-1)的细胞周期的影响;10)通过流式细胞仪分析上调ZNRD1对细胞(AGS、MKN28、NIH3T3)的凋亡的影响;11)通过MTT实验检测ZNRD1高/低表达对细胞(SGC7901、SGC7901/VCR、HL-60、HL-60/VCR)体外药物敏感性的影响;12)通过肾包膜下移植法检测ZNRD1高/低表达对细胞(SGC7901、SGC7901/VCR)体内药物敏感性的影响;13)通过流式细胞仪分析ZNRD1高/低表达对细胞(SGC7901、SGC7901/VCR、HL-60、HL-60/VCR)内阿霉素蓄积和泵出的影响;14)通过透射电镜检测上调ZNRD1对SGC7901细胞凋亡敏感性的影响;15)通过流式细胞仪和DNA梯度试验检测ZNRD1高/低表达对细胞(SGC7901、SGC7901/VCR、HL-60)凋亡敏感性的影响;16)通过基因芯片检测ZNRD1高/低表达对胃癌细胞内基因表达谱的影响;17)利用RT-PCR、Western blot对基因芯片的结果进行鉴定;18)利用报告基因实验检测ZNRD1对cyclin D1的启动子活性的调节作用;19)利用报告基因实验检测ZNRD1高/低表达对MDR1的启动子活性的调节作用;20)利用激酶试验检测ZNRD1对cyclin E-CDK2 激酶活力的影响;21)利用反义核酸技术抑制p21的表达;通过流式细胞仪检测抑制p21对ZNRD1介导的细胞周期阻滞的影响;22)利用反义核酸技术抑制p27的表达;通过流式细胞仪检测抑制p27对ZNRD1介导的细胞周期阻滞的影响;23)利用脂质体转染法上调Skp2的表达;通过流式细胞仪检测上调Skp2对ZNRD1介导的细胞周期阻滞的影响;24)利用Western blot检测ZNRD1对p27和Skp2的蛋白稳定性的影响;25)利用微血管密度实验检测ZNRD1对AGS、MKN28细胞裸鼠移植瘤微血管形成的影响;26)利用ELISA检测ZNRD1对AGS、MKN28细胞培养上清和移植瘤匀浆中VEGF165含量的影响;27)利用脂质体转染法、MTT实验、肾包膜下移植法、流式细胞仪和DNA梯度试验检测新耐药相关分子DARPP-32对细胞(SGC7901、SGC7901/VCR、对阿霉素耐药的胃癌细胞SGC7901/ADR)多药耐药表型的影响;利用脂质体转染法和MTT实验检测下调ZNRD1对DARPP-32介导的胃癌多药耐药的调控作用。

objective the effects of anthocyanin pigment from maize purple plant on resisting lipid peroxidation were investigated.methods the inhibition of anthocanin pigment from maize purple plant was examined in vitro that autcoxudation of lecithin liposome system induced by fe2+.50 mice were randomly divided into 5 groups.vehicle and different dose anthocyanin pigment from maize purple plant were given respectively,and then experimental injury of mice liver was induced to make use of bromobenzene and the content of mda was determined in liver homogenate.results the inhibition rates of anthocyanin pigment from maize purple plant on linoleic acid oxidation was higher that of ascorbic ascorbic acids.the content of mda in homogenate in middle and high dose of pigment were significantly lower than that in low dose and the injured control group.there were no significat differences in the content of mda in homogenate between low dose group and injury control group.conclusion the anthocyanin pigment from maize purple plant have the capability to resist lipid peroxide.

目的 探讨玉米紫色植株色素抗脂质过氧化的作用。方法体外实验测定玉米紫色植株花色苷色素在fe2+引发的卵磷脂脂质体体系中抗氧化活性。体内实验,取50只小鼠随机分成5组,分别给予溶媒和不同剂量的色素,然后采用溴代苯致实验性肝损伤,测定肝匀浆的丙二醛含量。结果在由fe2+引发的卵磷脂脂质体体系中玉米紫色植株色素对脂质过氧化有明显的抑制作用,抑制率随样品的浓度增高而增大,并且明显优于抗坏血酸。在溴代苯致小鼠实验性肝损伤模型实验中,中、高剂量组的丙二醛含量均低于损伤模型组。低剂量组和损伤模型组比较丙二醛含量差异无统计学意义。

The results manifest that the concentration of glycocholic acid has satisfactory linear correlation whith peak area, regression equation: Y= 39579X + 57823,R2=0.9984;The lowest detected concentration is 0.02μg/mL with S/N=3 ; Average recovery rate is 98.49%; RSD为0.52%~2.00%in day, RSD为0.74%~4.98%between days;The average recovery rate is 99.35%, RSD is 2.24% in the first team, and it is 100.87%, RSD is 4.73% in the second team at the stability test. When the concentration of glycocholic acid was detected in the liver, kidney and brain tissue, corresponding tissue concentration was calculated by preparing the standard curve with the glycocholic acid standard preparation in vitro, but the result can only reflect the relative chang law of the concentration of glycocholic acid in tissue but not in real tissue. The result of tissue stability experiment is consistent with that of the serum experiment.

结果表明,在0.15625~10.0μg/ml范围内血清中甘氨胆酸浓度与峰面积呈良好的线性关系,回归方程Y= 39579X + 57823,R2=0.9984;以信噪比S/N=3为标准,最低检测浓度为0.02μg/ml;平均回收率为98.49%;日内RSD为0.52%~2.00%,日间RSD为0.74%~4.98%;稳定性试验第一组的平均含量为99.35%,RSD为2.24%,第二组的平均含量为100.87%,RSD为4.73%;测定了肝、肾、脑组织中甘氨胆酸含量,采用体外甘氨胆酸标准品制备标准曲线的方法计算了相应的组织浓度,其结果仅反映组织中甘氨胆酸浓度的相对变化规律,不能体现真正的组织浓度值,但对组织样品的稳定性进行了较细致的考察,其结果和血样结果基本一致。

Degradation test in vitro was carried out in phosphate buffer solution (0.1M, pH7. 4) at 37 ℃. The buffer solution was changed daily. Degradation test in vivo was implanted the sample to subdermal in adult ICH rat in the scapular area lateral to the dorsal midline. At suitable time the samples were recovered. Molecular weight changes in surface layer and bulk of polymer sample were measured by GPC and weight loss was determined gravimetrically. It was found that the degradation behavior can be regulated by changing the composition of copolymers. The critical compositions from surface to bulk degradation behavior for PGCA, PLCA, PLMCA, PLDCA systems were 15-20, 20-30, 30- 40, 40 of mol percent GA or LA unit in copolymers, respectively. The degradation behavior of PGCA, PLCA, PLMCA, PLDCA systems were compared and analysed. Some factors influencing the degradation character, such as copolymer composition, hydrophobicity, crystallinity and enzyme affect etc. played important role.

体外实验中材料降解环境为37℃,0.1M,pH7.4磷酸缓冲溶液,每天换液,定期取样;体内实验中将聚合物试样埋置于ICH小白鼠背部肩胛骨皮下部位,定期处死小鼠,取样,将体内体外样品进行重量损失及试样内外层分子量变化测定,分析各聚合物试样降解行为特性,实验结果证明,改变共聚物组成,可以调节各聚合体系降解行为特性,对PGCA,PLCA,PLMCA,PLDCA共聚体系,交酯摩尔百分含量15-20%,20-30%,30-40%,40%分别为各体系内降解行为特性由表面降解型向本体降解型过渡的临界转折点,交酯含量较低的聚合物不同程度地表现表面降解行为特性,论文对各共聚体系体内外降解行为作了分析对照,例如共聚物组成对材料降解速度与降解行为的影响;生物体内酶对降解行为的影响;材料亲疏水性,聚合物链段结晶行为及碳酸酯结构对材料降解行为的影响等,得出交酯/环碳酸酯共聚体系降解行为一些共性和规律。

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Fancy gold-plated dangling earrings with facetted White Opal crystals.

花式镀金悬垂耳环与facetted白欧泊水晶。

This essay chooses the study aim from biology teachers in middle school in Shi Jiazhuang which tells us that most of the middle school biology teachers in Shi Jiazhuang have the"burnout", lower successfulness, individualize.

本文选取石家庄市初中生物教师作为研究对象,运用问卷调查的方法对石家庄市初中生物教师职业倦怠的现状进行调查,调查结果发现,石家庄市初中生物教师这一群体普遍存在职业倦怠,情感枯竭程度偏高,成就感偏低,去个性化程度最为严重。

In measurements of a day,generallyspeaking,the photosynthesis of birch in mesophytic habitat is better than that in xerophytichabitat(peak values are 12.8,10.33μmolCO2m-2s-1 respectively);that of sexual birch inmesophytic habitat is better than that of clone birch(peak values are 9.87,6.71μmolCO2m-2s-1respectively);that of young tree is better than that of seedling(peak values are12.37,10.05μmolCO2m-2s-1 respectively).

在一天中的各个时刻,总体说来,中生生境生长的白桦光合作用超过旱生生境生长的白桦光合作用(净光合速率峰值分别为12.8、10.33μmolCO2m-2s-1);白桦幼树的光合作用超过白桦幼苗(净光合速率峰值分别为12.37、10.05μmolCO2m-2s-1);中生生境有性白桦的光合作用超过无性白桦的光合作用(净光合速率峰值分别为9.87μmolCO2m-2s-1、6.71μmolCO2m-2s-1)。