细胞化学

Isolation, cultivation and identification of human embryonic NSC from cerebral cortex of 7-12 week abortus. By using flow cytometry and MTT assay, the effects of various concentration of TSPG and TSPG cooperating with cytokines in NSC culture media for 3 days on proliferation of human embryonic NSC has studied. By employing immunocytochemistry assay of the expression of tyrosine hydroxylase, the effect of different dilution of TSPG and TSPG cooperating with IL-1 on induced differentiation of human embryonic NSC into dopaminergic neuron has researched.

从7～12周人工流产胚胎脑组织中分离培养人胚胎NSC，免疫细胞化学鉴定NSC特异性nestin抗原表达，在NSC的培养体系中加入TSPG，采用流式细胞术和MTT法检测不同浓度TSPG以及TSPG与EGF，bFGF联合应用对人胚胎NSC增殖的影响；经免疫细胞化学检测培养细胞酪氨酸羟化酶表达，分析不同浓度TSPG以及TSPG与IL-1联合应用对NSC定向诱导为DA能神经元的影响。

The Journal of Histochemistry and Cytochemistry is a official journal of the Histochemical Society.

组织化学和细胞化学杂志》是该学会的官方期刊，它报道组织化学和细胞化学？？？

Cells were characterized by immunohistochemistry, cytochemistry, FACS and ultrastructure to identify and detect the differentiated population and markers.

通过免疫细胞化学、细胞化学、流式细胞分析法及透射电镜对其分化后细胞进行鉴定。

To isolate mononuclear cells from foetus umbilic cord blood, After MNCs were incubated in the presence of APS group (APS 200 mg·L-1) and control group for 24 h,the cells were stimulated with Epo（5 IU/mL） for 0,2,5,30 min; STAT5 was measured by ICC and laser confocal scanning microscope, JAK2、STAT5 in nucleus and cytoplasm were measured by western-blotting-ECL in each group at different groups.

分离纯化胎儿脐带血单个核细胞，在常规体外培养体系加入APS（200 mg·L-1）作用细胞24 h，促红细胞生成素分别刺激0，2，5，30 min，免疫细胞化学与激光共聚焦显微镜观察细胞STAT5的表达；Western-blotting-增强化学发光法检测细胞JAK2与浆和核中STAT5的表达。

Methods: A 5ml bone marrow was extracted from the lilac of human volunteers. By Percoll fluid and density gradient centrifugation, the MSC was obtained; after the cells filled the bottom of vessel, subcultured them, when they subculture in third generation, redigested them, 500 R/min centrifugate, alter the completed medium to chemical definition medium, examined the form change and prolifration of cells by invert microscope, toluidine blue stain、immunocytochemical stain and RT-PCR to test the type Ⅱ collagen mRNA and proteoglycan.

取健康成人髂后上棘处骨髓5ml，经percoll液分离后密度梯度离心，〓／ml密度接种培养，观察原代细胞的贴壁、增殖状况，细胞长满瓶底后进行传代培养；传至第三代细胞，重新消化后以500转／分钟轻度离心5分钟，〓／ml接种，改用化学限定培养基代替完全培养基培养，倒置显微镜观察细胞生长情况及形态变化，甲苯胺蓝染色观察诱导细胞合成细胞外基质中的蛋白多糖，免疫细胞化学染色检测ECM中Ⅱ胶原的蛋白合成，RT-PCR鉴定诱导细胞Ⅱ胶原mRNA的表达。

Part one Isolating, culturing, chondroblast induction and evaluation of human MSC in vitroObjective:To examine the property and proliferation ability of primary culturing MSC ,mvestgate the effect of lightly centrifugation and chemical definition medium to induce MSC differentiate into chondroblast.

结果：原代骨髓基质干细胞易于体外培养，增殖旺盛；经轻度离心和化学限定培养基诱导后细胞呈高密度生长，仍保持较旺盛的增殖能力，细胞转化成圆形肥大细胞，甲苯胺蓝染色显示软骨细胞外基质 v的特异性异染，11胶原蛋白免疫细胞化学染色呈强阳性，BT干CR鉴定显示＊胶原mRNA丰富的表达。

The aim are:lTo examine the proliferation ability and potential chondroblast differentiation ;2To find an ideal condition stimulated BMSC differentiate into chondroblast;3To examine the chondroblast proliferation in porous scaffolds and to explore the interaction between cells and materials;4To examine the release of cytokine in vitro.

取健康成人略后上棘处骨髓 5ml，经 percoll液分离后密度梯度离心，10'砌l密度接种培养，观察原代细胞的贴壁、增殖状况，细胞长满瓶底后进行传代培养；传至第三代细胞，重新消化后以500转／分钟轻度离心 5分钟，10V加 l接种，改用化学限定培养基代替完全培养基培养，倒置显微镜观察细胞生长情况及形态变化，甲苯胺蓝染色观察诱导细胞合成细胞外基质QCM）中的蛋白多糖，免疫细胞化学染色检测ECM中＊胶原的蛋白合成，RT干CR鉴定诱导细胞＊胶原mRNA的表达。

After the cell growth curves was recorded, RPE cells of the 3-5th passages were utilized. 2、Three different siRNA (siRNAl,siRNA2,siRNA3) targeting against human cx43 gene and one negative control siRNA were designed and transfected into cultured human RPE cells via liposome reagent. The most effective siRNA can be determined by semi-quantitative reverse transcription PCRRT-PCR. 3、To the most effective siRNA, after transfected into human RPEs with different concentration, the cellular proliferate activities were messured by MTT colorimetry ; the percentages of RPE in different cell circle phase was assayed by FCM; the changes of phenotypical properities were observed with SCM; the protein expression of cx43 was studied through immunocytochemistry stain and Weston blot; the communication intercellular was calculated with FRAP; and the ability of recovery was assessed by using an in vitro wound healing model.4、The total proteins of siRNA1 and RPE were seperated by two-dimensional gel electrophoresis and visualized by silver staining. Proteins with significant expression alterations were selected and their peptide mass fingerprints (PMFs were obtained by matrix-assisted laser desorption/ionization time of flying mass spectrometry (MALDI-TOF-MS).The PMFs were used to search NCBInr database by Auto MS-Fit software.

实验方法：1、培养原代的人RPE细胞，经过细胞角蛋白、S-100和神经胶质原纤维酸性蛋白免疫细胞化学鉴定后，通过AO/PI染色技术确定培养细胞的存活率，描记其生长曲线，第3-5代用于以下细胞实验2、生物合成针对人cx43基因的三条小干扰RNA和一条阴性RNA通过脂质体转染RPE细胞后，通过RT-PCR的方法确定抑制效率最高的干扰片断3、将该片段以不同浓度通过阳离子脂质体转染培养的人RPE细胞后，采用MTT法观察其对细胞的增殖力的作用；通过流式细胞仪观察其对细胞周期的影响；通过扫描电镜观察其对细胞形态的影响；通过免疫细胞化学和Weston blot观察其对cx43蛋白表达的作用；采用激光共聚焦和荧光淬灭恢复技术观察荧光恢复速率平均百分率，评价其对细胞间通讯功能的影响；通过制作RPE细胞损伤模型，观察其对损伤修复能力的作用4、分离纯化转染siRNA的RPE组和正常对照组RPE细胞的全部蛋白质，应用等电聚焦电泳和SDS-PAGE双向电泳技术，银染显示分离出的蛋白质斑点，经凝胶图像分析软件对两个样本进行胶图分析，寻找差异蛋白点。

METHODS: SGC7901 cells were transfected with siRNAs targeted against the human PLK1 by chemosynthesis. The PLK1 mRNA levels of cells transfected with siRNAs were monitored by real time PCR and the protein levels were detected by immunocytochemistry. Cell proliferation was evaluated by direct cell counting with trypan blue staining.

化学合成针对PLK1的小干扰RNA，转染至胃癌细胞株SGC7901中，利用实时定量PCR、免疫细胞化学、台盼蓝活性细胞计数和流式细胞术对PLK1的基因表达、细胞增殖、细胞周期和凋亡的检测。

The status of cell growth was observed under inverted microscope and the morphological characters were studied with the cells stained by Wrights staining .

倒置显微镜动态观察细胞生长情况，瑞氏染色观察细胞形态特征，并采用细胞化学和免疫细胞化学方法对获得的细胞进行鉴定。

I didn't watch TV last night, because it .

昨晚我没有看电视，因为电视机坏了。

Since this year, in a lot of villages of Beijing, TV of elevator liquid crystal was removed.

今年以来，在北京的很多小区里，电梯液晶电视被撤了下来。