Rapid determination of DNA synthesis in adherent cells grown in microtiter plates

A specific, rapid, and economical method for measuring the extent of DNA synthesis in adherent rat hepatoma H4-II-E cells grown in 96-well microtiter plates is described. The adherent cells were pulsed for 1 h with [methyl-3H]thymidine, released from the substratum by trypsinization, and collected on fiberglass filters with a MASH II cell harvester. The amount of radioactivity incorporated was directly proportional to the number of cells per well. Growth curves generated by measuring [methyl-3H]thymidine incorporation and counting the number of cells per well were identical. Experiments with inhibitors of DNA, protein, and RNA synthesis demonstrated that this method selectively measured DNA synthesis. In addition, [3H]thymidine uptake showed excellent correlation with autoradiographic assessment of DNA synthesis. This specific and sensitive method for determining DNA synthesis in microtiter cultures should facilitate studies of effects of various growth-controlling agents on epithelial, fibroblastic, and other cells which grow as adherent cells in culture.     

Epitope-relatedness and phenotyping of hepatic cytochromes P-450 with monoclonal antibodies.

The epitope-specific cytochrome P-450 content of animal livers was analysed by radioimmunoassay using a panel of seven monoclonal antibodies (MAbs) made to a 3-methylcholanthrene-induced rat liver cytochrome P-450. Competitive radioimmunoassays utilizing a reference radiolabelled MAb and a series of unlabelled MAbs indicated that there are at least three distinct classes of MAbs to different epitopes on cytochrome P-450. In addition, a direct radioimmunoassay employing a radiolabelled second antibody detected MAb-specific cytochromes P-450 in livers from different animals. This radioimmunoassay detected large elevations in the levels of these cytochromes P-450 in the livers of 3-methylcholanthrene-treated rats and C57BL/6 mice compared with untreated rats, 3-methylcholanthrene-treated DBA/2 mice or guinea pigs. The two complementary radioimmunoassay methods are sensitive, efficient, and easily applicable for screening large number of tissue samples for MAb-defined cytochrome P-450 phenotype.     

新城疫病毒单克隆抗体的特性及应用

建立了8个分泌抗新城疫病毒(NDV)特异性单克隆抗体(McAb)的杂交瘤细胞株,根据它们的免疫生物学特性可以分为三种类型:(1)具有FA和ELISA特性(FN1、FN4、FN29、FN30、FN35、FNl22);(2)具有FA、ELISA和HI特性(FN7);(3)具有ELISA、HI特性和中和能力(FN106),根据FN30和FN106的ELISA试验,可将11个NDV毒株分为二种不同的抗原群,应用FN4-FITC,FN7-FITC和FN29-HRP试剂,对人工感染NDV和野外送检病例检测结果表明,单抗试剂的DFA阳性率(92.3％)高于病毒分离阳性率(87.2％),两种方法的符合率89.7％,这些单抗试剂用于临床诊断敏感性和特异性高,且方法快速、简便。     

广谱肾综合征出血热病毒单克隆抗体的A35的生物学性状

具有中和及血凝抑制活性的、能和世界各地分离到的肾综合征出血热病毒(HFRSV)发生反应的、广谱的单克隆抗体(McAb),对HFRSV的诊断和分子生物学研究都有重要意义。 本文着重比较了HFRSV McAbA5、A19、A25-1、A25-7和A35的生物学性状,并观察了对感染动物的实验治疗效果。     

细胞色素C_3在Rhodopseudomonas caosulata载色体光合磷酸化中的作用

细胞色素C_3不仅能增进除去铁氧还蛋白载色体的循环光合磷酸化活性的恢复,而且也增进以DCPIPH_2为电子供体,维生素K_3、反丁烯二酸分别为电子受体构成的非循环光合磷酸化活性的恢复。 氩气相下,细胞色素C_3促进正常载色体光合磷酸化活性的最适浓度是1.8 μmol,当PMS存在时,这一促进作用随C_3浓度的增加而直线上升,然后呈稳态。 Antimycin A(10~(-7)mol/L)能充分抑制C_3参与的光合磷酸化活性,这一抑制现象在PMS存在时消失。 o~-phenanthroline(1×10~(-5)mol/L.)对C_3参与的光合磷酸化活性亦具抑制作用,并被PMS的添加而消失,当浓度提高时(10~(-3)mol/L),抑制现象不因PMS的存在而消失。 氢气相下的载色体光合磷酸化活性比氩气相下的低,并且随着载色体贮存(-20℃)时间的增长而急剧下降,24h贮存,丧失活性达60％。C_3对氢气相下的光合磷酸化活性具明显的促进作用,而铁氧还蛋白则不能,但两者同时存在时,其磷酸化活性显著提高。     

鸡胚视网膜发育中花生凝集素结合糖蛋白的图谱

我们应用微量(1—3微克蛋白)等电点聚焦和SDS双向电泳对鸡胚视网膜发育的第4天(E_4)到第16天(E_(16))总蛋白图谱进行分析,并结合印迹的PNA免疫酶标反应,分析了总蛋白中与PNA结合的糖蛋白图谱的变化。E_4时没有看到PNA结合糖蛋白的任何阳性反应的斑点,E_6时这种糖蛋白的种数突然增加,约占总蛋白数的20%。新的糖蛋白增加的速率在E_6时最快,存其后的发育阶段(E_8—E_(16))慢于总蛋白增加的速率。这种糖蛋白大都是等电点在中性pH范围内,中等大小分子量的糖肽。视神经的总蛋白中含PNA结合糖蛋白数的比例高于同一发育年龄视网膜中的比例,这说明糖蛋白可能主要存在于质膜上。视网膜的总蛋白图谱相似于前脑的,但与视顶盖的更相近。本文对这种能与PNA结合,即含有D-半乳糖残基的糖蛋白在视网膜发育中的意义进行了讨论。     

广东省蝗虫二新种(直翅目:蝗总科)

在整理广东省蝗虫标本时,发现二新种。模式标本保存于中山大学昆虫学研究所标本室。 1.蒲氏板齿蝗Sinstauchira pui新种(图1—9) 雄虫 体中型。头短于前胸背板,头顶平,顶端圆形,宽略大于长。触角丝状,到达后足股节基部,中段一节长为宽的2倍。颜面倾斜,具粗刻点,颜面隆起明显,中眼以上侧缘近平行,中眼以下逐渐展开,纵沟浅。复眼卵圆形,纵径为横径的1.38倍,为眼下沟的2.57倍。眼间距为触角间颜面隆起的1.6倍。前胸背板前缘较平,后缘呈弧形突出;中隆线明     

Distribution and biosynthesis of aminopeptidase N and dipeptidyl aminopeptidase IV in rat small intestine

The regional, cellular and subcellular distribution patterns of aminopeptidase N and dipeptidyl aminopeptidase IV were examined in rat small intestine. Aminopeptidase N of brush border membrane had maximal activity in the upper and middle intestine, while dipeptidyl aminopeptidase IV had a more uniform distribution profile with relatively high activity in the ileum. Along the villus and crypt cell gradient, the activity of both enzymes was maximally expressed in the mid-villus cells. However there was substantial dipeptidyl aminopeptidase IV activity in the crypt cells. Both enzymes were primarily associated with brush border membranes in all segments, however, in the proximal intestine, a significant amount of dipeptidyl aminopeptidase IV activity was associated with the cytosol fraction. The cytosol and brush border membrane forms of dipeptidyl aminopeptidase IV were immunologically identical and had the same electrophoretic mobility on disc gels. In contrast, the soluble and brush border membrane-bound forms of aminopeptidase N were immunologically distinct. When the total amount of aminopeptidase N and dipeptidyl aminopeptidase IV was determined by competitive radioimmunoassay, there were no regional or cellular differences in specific activity (enzyme activity/mg of enzyme protein) of either enzyme in brush border membrane and homogenate. The specific activity of both enzymes in a purified Golgi membrane fraction as measured by radioimmunoassay was about half that of the brush border membrane fraction. These results suggest that (1) aminopeptidase N and dipeptidyl aminopeptidase IV have different regional, cellular and subcellular distribution patterns; (2) there are enzymatically inactive forms of both enzymes present in a constant proportion to active molecules and that (3) a two-fold activation of precursor enzyme forms occurs during transfer from the Golgi membranes to the brush border membranes.     

Preliminary crystallographic studies of lobster D-glyceraldehyde-3-phosphate dehydrogenase and the modified enzyme carrying the fluorescent derivative

When the active-site carboxymethylated D-glyceraldehyde-3-phosphate dehydrogenase is irradiated with ultraviolet light in the presence of NAD+, a fluorescent NAD derivative that is covalently linked to the enzyme is obtained. A preliminary crystallographic study of this fluorescent derivative, as well as of the native and the carboxymethylated enzymes from Palinurus versicolor, showed that they are isomorphous and belong to space group C2 as reported for the native enzyme from Palinurus vulgaris. The three forms of the enzyme, although they have identical unit cell parameters, differ considerably in their diffraction patterns, indicating marked differences in conformation in spite of the fact that they differ chemically only in a restricted region around the active site.     

广西贵县的螟害规律及其预测方法的探讨

1.从广西贵县1958—1964年的七年螟害调查资料,可以将全年螟害的消长趋势,划分为世代递减发生型、世代递增发生型和第三世代多发型等三种螟害发生型。 2.形成三种螟害发生型的原因,主要系由于水稻播植期的早晚与螟虫发生为害的不同组合所致。水稻播植期较早的年份,形成世代递减发生型;播植期较晚的年份,形成世代递增发生型:播植期居中的年份,则形成第三世代多发型。 3.本文根据历史资料,订出早、中、晚播植期的标准,以此标准为坐标图上的横轴,各世代螟害率为纵轴,作出了播种期与螟害型的关系模式图。根据这个模式图,在某年掌握了早稻播种期以后,就可以作出该年螟害发生发展趋势的展望。 4.求算三种螟害发生型各世代之间的螟害增减比值历年平均数,在某年获悉早稻播种期和第一世代实测螟害率以后,就可以根据这个比值平均数,依次推算以后各世代的预测螟害率。 5.关于各世代螟害率的近期预测,本文主张以幼虫虫口密度为依据,求算虫口密度与其所造成螟害率的二次函数曲线方程式,在获得幼虫密度以后,代入方程式来计算近期预测的螟害率。