CONCENTRATION AND PURIFICATION OF BACTERIOPHAGE

1. A method for isolating a nucleoprotein from lysed staphylococci culture is described. 2. It is homogeneous in the ultracentrifuge and has a sedimentation constant of 650 x 10^–13 cm. dyne^–1 sec.^–1, corresponding to a molecular weight of about 300,000,000. 3. The diffusion coefficient varies from about 0.001 cm.²/day in solutions containing more than 0.1 mg. protein/ml. to 0.02 in solutions containing less than 0.001 mg. protein/ml. The rate of sedimentation also decreases as the concentration decreases. It is suggested, therefore, that this protein exists in various sized molecules of from 500,000–300,000,000 molecular weight, the proportion of small molecules increasing as the concentration decreases. 4. This protein is very unstable and is denatured by acidity greater than pH 5.0, by temperature over 50°C. for 5 minutes. It is digested by chymo-trypsin but not by trypsin. 5. The loss in activity by heat, acid, and chymo-trypsin digestion is roughly proportional to the amount of denatured protein formed under these conditions. 6. The rate of diffusion of the protein is the same as that of the active agent. 7. The rate of sedimentation of the protein is the same as that of the active agent. 8. The loss in activity when susceptible living or dead bacteria are added to a solution of the protein is proportional to the loss in protein from the solution. Non-susceptible bacteria remove neither protein nor activity. 9. The relative ultraviolet light absorption, as determined directly, agrees with that calculated from Gates'' inactivation experiments in the range of 2500–3000 Å. u. but is somewhat greater in the range of 2000–2500 Å. u. 10. Solubility determinations showed that most of the preparations contained at least two proteins, one being probably the denatured form of the other. Two preparations were obtained, however, which had about twice the specific activity of the earlier ones and which gave a solubility curve approximating that of a pure substance. 11. It is suggested that the formation of phage may be more simply explained by analogy with the autocatalytic formation of pepsin and trypsin than by analogy with the far more complicated system of living organisms.     

PURIFICATION AND CONCENTRATION OF ENTEROKINASE

A concentrated solution of purified enterokinase is conveniently prepared from the fluid contents of pigs'' duodena by means of fractional precipitation with ammonium sulfate under the proper pH conditions.     

A NEW RAPID TECHNIQUE FOR SALMONELLA ANTIGEN DETECTION USING A PARTICLE CONCENTRATION FLUORESCENCE IMMUNOASSAY (PCFIA)

A fluorescent assay employing polystyrene beads coated with antibody to a common structural antigen (CSA) of salmonellae has been developed to detect the presence of Salmonella. This sensitive technique is very fast, relatively simple and with further testing may be suitable for screening food products as well as clinical diagnostics. Comparison was made with a commercially available Salmonella antigen capture enzyme immunoassay (EIA). As little as one single colony-forming unit (CFU) could be detected by fluorescence after only 16 h of enrichment. the procedure requires only 1 h to run and uses proportionately smaller volumes of sample and reagents than EIA.     

A PROCEDURE FOR THE PURIFICATION OF BETA-LACTOGLOBULIN FROM BOVINE MILK USING GEL FILTRATION CHROMATOGRAPHY AT LOW pH

A simple and rapid procedure for the purification of beta-lactoglobulin (β-LG) from bovine milk is described. The procedure exploits the major difference in molecular mass of β-LG and other whey components and the existence of the former in monomeric form at acidic pH. Gel filtration of whey was carried out using a Bio-Gel P10 column at pH 3.0. Residual caseins and other milk proteins were excluded from the gel and β-LG and alpha-lactalbumin (α-LA) emerged as two fully resolved peaks. Sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) suggested that β-LG was purified to apparent homogeneity, while absorption, fluorescence, and circular dichroism spectroscopy indicated the native-like conformation of the protein. Western blot analysis revealed that the antibodies raised against the purified β-LG in rabbits also readily react with the commercial bovine protein. This procedure requires only 4–5 hr for the purification of about 10 mg of β-LG from a single run while using a small column (2.3 cm × 83 cm) of Bio-Gel P10 and has the potential for scaling up.     

一种嗜热厌氧纤维素降解细菌的分离纯化方法

根据纤维素降解细菌对不溶性纤维素底部的粘附作用,利用Ｈｕｎｇａｔｅ厌氧操作技术直接以不溶性纤维素粉为基质进行滚管,分离和纯化获得嗜热厌氧纤维素降解细菌。     

人工湿地净化污水处理厂低浓度尾水的效果

文章构建一种以双向横流过滤单元为核心的人工湿地系统,以实际污水处理厂尾水为进水,研究了尾水湿地系统净化效果,发现采用该湿地模式的出水能稳定提升1个等级。为了进一步探究湿地系统除污机理,以污水处理厂实际低浓度尾水为进水,研究了该湿地系统不同植物单元污染物去除功效,讨论了植物、基质与微生物的协同作用机制。研究结果表明:(1)在不同植物的去污效果对比实验中,花叶芦竹植物单元对COD、TN、NH₄⁺-N和TP的平均去除率最高,分别为20.11%、17.17%、28.08%和18.12%;(2)不同运行阶段时基质进出水端主要微生物群落差异较小;(3)系统反向进水时,湿地系统TN去除率可提升50.00%。研究结果对于尾水湿地建设具有一定指导意义。     

诱导凋亡:脊髓灰质炎病毒致细胞病变的机制

用脊髓灰质炎病毒(Poliovirus)减毒活疫苗株(中Ⅲ-2)感染人二倍体胚肺成纤维细胞株(KMBl7)后,细胞形态变化分为两个阶段第一阶段是致CPE过程,导致形态学上特有的细胞圆缩、体积缩小等CPE特征,经光学显微镜、荧光显微镜、细胞流式仪、电子显微镜、DNA凝胶电泳分析,证明该疫苗株诱导的细胞病理改变具典型的凋亡特征细胞圆缩、细胞核浓缩破裂、核染色质凝缩后分布于核膜边缘、绝大部分细胞出现在凋亡区域、DNA断裂。表明脊髓灰质炎病毒诱导的CPE本质是一个诱导细胞凋亡过程;第二阶段是Poliovirus诱导细胞坏死。进一步提示有可能通过抑制细胞凋亡提高脊髓灰质炎减毒活疫苗的产量。     

A RAPID, HIGHLY EFFICIENT METHOD FOR COLLECTING, FIXING, AND EMBEDDING PLANKTONIC AND OTHER SMALL CELLS FOR ELECTRON MICROSCOPY

A rapid, highly efficient method is described for fixation, dehydration, and embedding of small (e.g. planktonic) cells dispersed in large volumes of culture medium. The entire protocol, based on continuous filtration, can be completed within about an hour, and the yield of cells is very high. Fixation quality has been excellent with several different types of samples.     

A METHOD FOR THE FRACTIONATION AND PURIFICATION OF SEA URCHIN SPERM

Procedures for fractionation and purification of sea urchin sperm subunits were studied. Fragmentation of cells was observed rather than fractionation of cell subunits when classic sonication or homogenization techniques were used. Calcium shock in alcohol solution which successfully removes Protozoa cilia was not effective in removing sperm flagella. Alcohol was found to induce cohesion among whole sperm rendering separation of isolated subunits by centrifugation impossible. Calcium shock without alcohol in the medium was not effective in achieving dissociation of sperm subunits.
Successful fractionation of sperm subunits was achieved by application of a stepwise vortex-shearing action in a 0.22 micron filtered calcium-free medium. Analysis by Nomarski optics indicated little or no cellular fragmentation. Centrifugation with various discontinuous sucrose-sea water solutions proved to be a successful technique to purify the fractionated heads, midpieces, and tails.
The procedures reported here can be used to obtain fractionated and purified sperm cell heads, midpieces, and tails. These procedures do not require enzyme, homogenization, or sonication treatment of the material.     

A MODIFIED PROCEDURE FOR ISOLATION OF ASTROCYTE- AND NEURON-ENRICHED FRACTIONS FROM RAT BRAIN

Our previously published method for isolation of neurons with extensive processes (Farooq et al., 1977) has been modified to permit the isolation of both astrocyte- and neuron-enriched fractions. Rat cerebral tissue is incubated with acetylated trypsin and disrupted. The cell suspension is separated first by differential centrifugation and then by gradient centrifugation on discontinuous Ficoll gradients. The method is reproducible and is applicable equally well to immature and adult animals. The yield of astrocytes of 57% particle purity, and higher weight purity, is 4–7 × 10⁶ cells/brain, amounting to 1.5–2.0 mg of protein. The astrocytes appear to be a mixture of fibrous and protoplasmic types. The yield of neurons of 90% particle purity is 10–14 × 10⁶ cells/brain, amounting to 2.4–3.0 mg of protein. A total yield of neurons of 28–37 × 10⁶ cells/brain can be obtained at 70% purity. These preparations have been characterized by light microscopy and protein, RNA and DNA content.     

PROCEDURE FOR SELECTING STARTING CONFORMATIONS FOR ENERGY MINIMIZATION OF NUCLEOSIDES AND NUCLEOTIDES

ABSTRACT

The purpose of this study was to carry out a thorough search of the conformational space of various adenine-containing nucleotides, applying a previously published searching procedure, known as the representative method. This method, which reduces the number of starting conformations required to explore all the important regions of conformational space, appears to be successful in finding all (or nearly all) the putative low-energy conformations of each molecule.     

一种快速有效纯化DNA序列分析模板的方法

介绍一种ＤＮＡ序列分析模板的快速、有效的纯化方法。该法对ＤＮＡ模板的回收率可达９５％以上。多次测序结果表明,此法与其他常规纯化方法相比,具有简便、快速、有效、可靠等优点,其测序结果电泳带清晰,无模糊带及“鬼带”出现,重复性及稳定性较好。