钙调素化学发光标记的研究

用发光物质N-氨基丁基N-乙基异鲁米诺(ABEI),采用碳二亚胺缩合法,成功地标记了动物及植物钙调素(CaM),标记率为[ABEI]/[CaM]≈0.8-0.9.对ABEI标记CaM的反应条件、结合物的质量和保存时间进行了观察.此标记物稳定,-20℃可保存10个月以上,适用于化学发光免疫法测定生物样品中CaM含量.     

磷脂模型膜的时间分辨纳秒荧光光谱的研究

叙述了用时间分辨率纳秒荧光技术研究大豆磷脂或合成磷脂模型膜（脂质体）的物理状态变化与荧光寿命以及时间分辨各向异性测量参数变化相关性的实验结果。用荧光探剂ＭＣ５４０标记模型膜的结果表明,荧光寿命（τ）的变化与脂质／探剂比例、磷脂组成、磷脂的极性头部、脂肪酸酰链长度等有关。用ＤＰＨ（１,６－ｄｉｐｈｅｎｙｌ－１,３,５－ｈｅｘａｔｒｉｅｎｅ）标记磷脂模型膜的时间分辨各向异性分析结果表明,序参数（Ｓ）、     

Non-ependymal cilia in the habenulae and the interpeduncular nucleus of the frog tadpole

Summary Cilia of the 9+2 pattern are found electron microscopically in nonependymal cells of the habenulae and the interpeduncular nucleus of the tadpole of Rana esculenta at an early stage of development (8 mm length, head to tip of tail). A comparison is made between these and the ependymal and sensory cilia in the same specimens. The cilia project into the neuropil emerging from a perikaryon rich in free ribosomes and displaying a prominent Golgi apparatus. These perikarya contain dense core vesicles. Synapses with vesicles of the clear spherical type have been observed along the ciliary shaft. On a purely morphologic basis the authors hypothesize that these cilia, at least in this early ontogenetic stage, may extend considerably the conducting surface of the cell and represent a sensory structure which could be stimulated by terminal processes belonging to distantly located cells. In addition, they could also be involved in the trophic exchange of material with the adjacent structures.     

Parasites,predators and other arthropods associated withMusca domestica and other flies breeding in bovine manure

The species of flies breeding in bovine manure, their parasites and predators as well as other associated arthropods in 3 localities in and near Bangalore are recoreded. The flies areMusca domestica L.,Musca pattoni Austen,Stomoxys calcitrans L.,Physiphora aenea F.,Physiphora demandata F.,Sargus metallinus F.,Sepsis thoracica R.-D.,Sepsis nitens Wiedemann,Sphaerocera scabricula Hal, andLeptocera (Coproica) hirtula Rondani. Four species of hymenopterous pupal parasites of these flies have been obtained:Spalangia cameroni Perkins from pupae ofM. domestica, M. pattoni, S. calcitrans, P. aenea andP. demandata; Spalangia endius Walker from pupae ofM. domestica, M. pattoni andS. calcitrans; Spalangia nigroaenea Curtis from pupae ofM. domestica andS. calcitrans; andDirhinus trichiophthalmus Masi from pupae ofSargus metallinus F. Percentage parasitism of fly puparia in field samples was noted. Four species ofHister andAleochara puberula Klug feeding on various immature stages ofM. domestica, 2 species of scarabaeid beetles, 2 species of ants and a pseudoscorpion were also found in the manure. The importance of biotic regulatory factors in the control of flies is discussed.

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Résumé Dans 3 localités de Bangalore ou proches de cette ville, on a inventorié les espèces de mouches vivant dans le fumier de vache, leurs parasites et prédateurs ainsi que d'autres arthropodes associés. Les mouches récoltées sont:Musca domestica L.,Musca pattoni Austen,Stomoxys calcitrans L.,Physiphora aenea F.,Physiphora demandata F.,Sargus metallinus F.,Sepsis thoracica R-D.,Sepsis nitens Wiedemann,Sphaerocera scabricula Hal. etLeptocera (Coproica) hirtula Rondani. Quatre espèces d'hyménoptères parasites des pupes ont été obtenues de ces mouches:Spalangia cameroni Perkins deM. domestica, M. pattoni, S. calcitrans, P. aenea etP. demandata: Spalangia endius Walker deM. domestica, M. pattoni etS. calcitrans, P. aenea etP. demandata; Spalangia endius Walker trans; etDirhinus trichiophthalmus Masi deSargus metallinus F. On a noté le taux de parasitisme des nymphes dans les conditions naturelles. Quatre espèces deHister etAleochara puberula Klug s'attaquant aux divers stades non imaginaux deM. domestica, 2 espèces de scarabeides, 2 espèces de fourmis et un pseudoscorpion ont été également trouvés dans le fumier. L'importance de ces facteurs biotiques dans la régulation des populations de mouches est discutée.

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Paper presented at the seminar on utilization of farm wastes for rural industrial growth, National Dairy Research Institute, Bangalore, 31 st December, 1975.

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This paper is published with the permission of the Director-General, Indian Council of Medical Research, New Delhi     

壳聚糖固定化木瓜蛋白酶的研究

以壳聚糖为载体,成二醛为交联剂将木瓜蛋白酶固定化。5％戊二醛在4-6℃下处理载体5h,加酶液(3.5mg/mL蛋白,pH7.2)固定12h,活力回收达32％,作用于酪蛋白的半衰期为36天,其表观K_m(酪蛋白)值为0.075％(W/V),溶液酶的K_m值为0.086％;最适pH7.0～7.5,溶液酶为7.0～8.5。固定化酶在pH8.5以下,溶液酶在9.0以下活力稳定。固定化酶在45℃以下,溶液酶在75℃以下稳定。用6mol/L脲洗脱固定化酶4次(5.5h)活力仍有54.5％。用固定化酶处理啤酒浊度比对照下降了1.5-3.7倍,蛋白质含量下降了44％,冷藏(4℃)120天无冷混浊现象发生并保持了啤酒原有风味和理化性状。     

Halichondrin B and homohalichondrin B, marine natural products binding in the vinca domain of tubulin. Discovery of tubulin-based mechanism of action by analysis of differential cytotoxicity data

Data generated in the new National Cancer Institute drug evaluation program, which is based on inhibition of cell growth in 60 human tumor cell lines, were used to compare new compounds with agents of known mechanism of action in terms of their differential cytotoxicity. Two marine natural products, halichondrin B and homohalichondrin B, appeared repeatedly when the data base was probed with known antimitotic agents. We confirmed that both compounds were highly cytotoxic (IC50 values for L1210 murine leukemia cells of 0.3 and 1 nM, respectively), with accumulation of cells arrested in mitosis at toxic concentrations, that both inhibited the polymerization of purified tubulin, and that both inhibited microtubule assembly dependent on microtubule-associated proteins. Limited amounts of homohalichondrin B, the less active agent, were available, so only halichondrin B was studied in detail. Halichondrin B did not interfere with colchicine binding to tubulin, but it was a noncompetitive inhibitor of the binding of vinblastine to tubulin (apparent Ki, 5.0 microM). Halichondrin B was therefore compared with other agents which interfere with the binding of vinca alkaloids to tubulin (vinblastine, maytansine, dolastatin 10, phomopsin A, rhizoxin) in terms of its effects on tubulin polymerization, inhibition of GTP hydrolysis, inhibition of nucleotide exchange, and stabilization of tubulin, as well as the quantitative assessment of its effects on vinca alkaloid binding and inhibition of cell growth. Since halichondrin B was originally isolated from the same organism as the phosphatase inhibitor okadaic acid, and since it is about 50-fold more effective than okadaic acid as an inhibitor of L1210 cell growth, perturbations of cellular microtubules observed following treatment with okadaic acid should be interpreted cautiously.     

Thrombomodulin as a marker of radiation-induced endothelial cell injury.

Cultured confluent human umbilical vein endothelial cells were irradiated in vitro with 60Co gamma rays at doses from 0 to 50 Gy. After irradiation thrombomodulin was measured at different times over 6 days in the supernatants of endothelial cell culture medium, on the surface of the cells, and within the cells. At 24 h after irradiation, an increase in the release of thrombomodulin from irradiated endothelial cells and an increase in the number of molecules and the activity of thrombomodulin on the surface of the cells were observed; these reactions were dependent on radiation dose. The capacity of the cells to produce and release thrombomodulin was decreased from 2 to 6 days after exposure to 60Co gamma rays. Our data indicate that radiation can injure endothelial cells, and that thrombomodulin may be used as a marker of radiation-induced injury in endothelial cells. The interrelationship between the dysfunction of irradiated endothelial cells and the pathological mechanisms of acute radiation disease is also discussed.     

Separation of dissociated thyroid follicular and parfollicular cells: association of serotonin binding protein with parafollicular cells

Parafollicular cells (PC) of the sheep thyroid gland are neural crest derivatives that synthesize and release the biogenic amine serotonin (5-HT) as well as the hormone calcitonin. The thyroid also contains a highly specific serotonin-binding protein (SBP). Separation of dissociated thyroid cells was done to study the cellular localization of SBP and to develop a means of isolating PC for study. Various methods were used to obtain an enriched and purified population of PC. Minced thyroid glands were enzymatically dissociated and the cells were layered on a Ficoll linear density gradient. Fractions obtained from the gradient were examined for cell number, viability, 5-HT concentration, SBP activity, and morphology by electron microscopy. One of the fractions was found to be enriched in PC. High levels of 5-HT and SBP were also found in this fraction, whereas these levels were low where the majority of cells were found. This PC-rich fraction, however, contained numerous follicular cells (FC); therefore, additional approaches to cell separation were used. FC can be stimulated in vitro with thyroid stimulating hormone (TSH) to become intensely phagocytic. When stimulated cells were incubated in the presence of silica microspheres, the FC engulfed the microspheres, which were toxic to them. PC did not become phagocytic and were unharmed by the microspheres. Suspended cells, after incubation with microspheres, were centrifuged on a discontinuous gradient, and a PC-rich fraction was obtained. Silica, however, interfered with analysis of SBP. Another method to take advantage of the phagocytic potential of FC was therefore used. TSH-stimulated cell suspensions were passed through a column of sepharose to which thyroglobulin had been coupled. Stimulated FC apparently adhered to the beads and were retained by the columns. Fractions eluting from the columns were greatly enriched with PC. These fractions contained high levels of 5-HT and SBP, and considerably reduced FC contamination was found by quantitative electron microscopy. It is concluded that SBP is localized to PC in the sheep thyroid. The idea that these cells resemble serotonergic neurons in their mechanisms of 5-HT storage is supported.