Two-Hour Embedding Procedure for Intracellular Detection of Viruses by Electron Microscopy

An embedding method requiring only 2 h to complete and giving excellent ultrastructural preservation has been used for the rapid detection of viruses in tissue cultures. The method has also been applied successfully to mammalian tissue. It provides a rapid technique for identifying viruses isolated in tissue cultures, for screening cultures for adventitious agents, and for examining tissue biopsies for viruses.     

Semi-continuous ethanolic fermentation using a novel yeast settling and recycle technique

Summary A novel technique for the rapid settling of yeast cells is outlined. An inert, high density powder is added to a yeast suspension and the pH of the suspension is switched rapidly from 4.5 (fermentation pH) to 8.0. Large, rapid settling flocs of yeast are formed immediately. This technique has been applied to the recycling of yeast from an ethanolic fermentation.     

应用PCR技术检测串珠镰刀菌

串珠镰刀菌(Fusarium moniliform)是一种危害严重、在世界各地广泛流行的植物病原真菌,当前对串珠镰刀菌的鉴定主要根据其菌丝体及再生菌丝的形态结构学特征及染病作物的病害症状来进行鉴定。这些鉴定方法相对简单并在很大程度上依赖于经验,受主观因素影响较大。采用串珠镰刀菌种特异性的寡聚核苷酸为引物,运用PCR技术对串珠镶刀菌进行检测是一种快速可靠的检测鉴定方法,它无需病原菌的分离培养纯化,能从感病的玉米组织中直接实现对串珠镰刀菌的快速检测。经对霉变玉米样品和玉米穗腐病组织的检测,证明该方法是一种快建、有效的方法,具有重要的实际应用价值。     

Differentiation of Virus Types in Foot-and-Mouth Disease (FMD)

An improved complement-fixation technique in foot-and-mouth disease is described in details. In the development of the technique consideration has been given to the established presence of an antigen-excessphenomenon as well as to the linear relationship between amounts of immune serum used and complement fixed. The increased accuracy of the technique has made it possible more precise and rapid to detect even small serological differences.     

PCR-SSCP技术在微生物检测中的应用

聚合酶链式反应-单链构象多态性(PCR-SSCP)是一种能够检测DNA突变的分子生物学分析技术,具有快速、简便、灵敏与适于大样本筛选的特点,近年来被广泛应用于生命科学领域的研究。对PCR-SSCP技术在微生物检测研究领域的应用和发展作简要的介绍。     

PCR-SSCP--rapid and easy detection of DNA-sequence changes]

PCR-SSCP analysis is a rapid, simple and sensitive technique for detection of various mutations, including single nucleotide substitutions, insertions and deletions, in PCR-amplified DNA fragments. Here I review, the principle and sensitivity of the technique, and also show how this technique has been used in clinical and basic medical sciences.     

Synthesis of the human insulin gene. Part II. Further improvements in the modified phosphotriester method and the synthesis of seventeen deoxyribooligonucleotide fragments constituting human insulin chains B and mini-CDNA.

The purification of protected deoxyribooligonucleotides containing phosphotriester internucleotidic linkages has been improved by developing a deactivated silica gel chromatographic technique. The efficiency of this technique as applied in the modified phosphotriester approach has been demonstrated in the rapid synthesis of seventeen pure fragments constituting the sequence of human insulin B and mini-C DNA. The sequence of each oligomer was confirmed by the two-dimensional mobility shift method of fingerprinting.     

A rapid mixing-photocrosslinking technique to study the dynamics of nucleic acid-protein interactions

A rapid mixing-photocrosslinking technique has been developed to investigate the kinetics of protein-nucleic acid interactions. With this technique, binding of nucleic acid to protein is first synchronized by rapid mixing in a stopped-flow apparatus. The intermediates formed at different stages of the binding process are then "frozen" by photocrosslinking with a 10-microseconds uv light pulse at various times after mixing. By analyzing structural changes of these intermediates as a function of time, one can obtain the information concerning the dynamic aspects of the interaction. This technique may also be applied to other macromolecular interactions in biological systems.     

Advanced short-lived nuclide NAA with application in the life sciences

Biological Trace Element Research - A new technique for short-lived nuclide activation analysis has been developed that compensates the rapid radioactive decay during the counting period by...     

Detection of depolymerase isoenzymes after electrophoresis or electrofocusing, or in titration curves

The cup-plate technique makes it possible to detect enzyme activities after diffusion into buffered, substrate-containing agar gels. This technique has been used after nondenaturing blotting transfer in order to detect depolymerizing enzyme activities once analytical protein separation (e.g., by electrophoresis, electrofocusing, or titration curves) has been completed. This rapid and very sensitive method was successfully applied to the enzymes polygalacturonate lyase, polygalacturonate hydrolase, endoglucanase, and xylan hydrolase. Other possible applications are presented.     

Direct visualization of collagens and procollagens in polyacrylamide gels.

This report describes a procedure that results in the rapid visual identification of collagens and procollagens in polyacrylamide gels. The technique results in a pink stain for collagenous proteins and a blue stain for all other proteins. The color difference has been evaluated spectrophotometrically. The absorbance maxima for collagen-Coomassie blue R250 complexes in gels is 520–535 nm, and the maxima for all other protein-Coomassie blue R250 complexes that we tested is 550–560 nm. This technique will facilitate the identification of collagenous proteins in complex mixtures of proteins derived from cell membranes, whole cell extracts, conditioned media, and extracellular matrices. We use the technique to detect procollagens in human diploid fibroblast conditioned media. The technique is simple, relatively rapid, has utility for proteins extracted by a variety of methods, and is applicable to all polyacrylamide gel systems in general use.     

A rapid chitin synthase preparation for the assay of potential fungicides and insecticides

Summary A rapid technique for the preparation of large quantities of solubilized chitin synthase is described. The enzyme derived is very stable and has high specific and total activities. It has been used to screen compounds with potential fungicidal and insecticidal properties.     

Synapsin I from human brain. Phosphorylation by Ca2+, phospholipid-dependent protein kinase

Synapsin I has been isolated from human brain by a rapid and efficient purification technique, and its phosphorylation by human brain Ca2+, phospholipid-dependent protein kinase (protein kinase C) has been studied. The inhibitory effect of calmodulin on this process has been demonstrated. It is also found that non-esterified fatty acids and acidic phospholipids are inhibitory for synapsin I phosphorylation by Ca2+, calmodulin-dependent protein kinase II.     

A simple technique for the measurement of swimming speed of Chlamydomonas

A simple technique for the measurement of swimming speed of Chlamydomonas reinhardi is described. The technique is based on a rapid statistical counting method which treats the swimming cells as a dilute, kinetic gas and its accuracy has been confirmed by direct photomicrographic measurements. In principle, this technique can be applied to any suspension of organisms satisfying certain statistical criteria.     

The use of a simple enzyme assay in ‘seed-hardening’ studies

A simple technique for the assay of the enzyme, catalase is described. The method has been used to show that vegetable seeds that had been submitted to pre-sowing ‘hardening’ cycles of imbibition and drying, as well as having more rapid germination than controls, had greater catalase activity.     

Incremental analysis: The application to quantitation of both enzyme activity and inhibitory activity in complex subcellular fractions

Single stage conventional analysis of neutral protease activity present in certain tumour cell cytoplasmic fractions has been demonstrated to be totally misleading due to hidden errors introduced by the complex interactions of the protease with a cytoplasmic inhibitor of this enzyme. A novel system has been developed which enables the rapid simultaneous quantitative analysis of neutral protease activity and inhibitor activity. This technique employs an insoluble fluorescein-labelled substrate (polymeric collagen fibrils) and relies on the sensitive fluorimetric assay of solubilised fluorescein-labelled peptides. This technique has been termed “incremental analysis” and the advantages of incremental analysis over conventional analysis methods have been described in detail. The experimentally obtained results can readily be resolved graphically or by simple computation.     

Qualitative PCR-ELISA protocol for the detection and typing of viral genomes

PCR is an established technique providing rapid and highly productive amplification of specific DNA sequences. The demand for equally rapid, sensitive and objective methods to achieve detection of PCR products has led to the coupling of PCR with ELISA. PCR-ELISA involves direct incorporation of labeled nucleotides in amplicons during PCR-amplification, their hybridization to specific probes and hybrid capture-immunoassay in microtiter wells. PCR-ELISA is performed in 1 d and is very flexible, with the ability to process simultaneously up to 96 or 384 samples. This technique is potentially automatable and does not require expensive equipment, and thus can be fundamental in laboratories without access to a real-time PCR thermocycler. PCR-ELISA has mainly been used to detect infectious agents, including viruses, bacteria, protozoa and fungi. A PCR-ELISA protocol for the qualitative detection of papillomavirus genomes and simultaneous typing of different genotypes are detailed here as an example of the technique.     

The enantioselectivity of enzymes monitored using a chiral detector

Summary A chiral detector coupled to an HPLC system has been used to follow the enantioselectivity of the reduction of bicyclic ketones catalysed by dehydrogenase enzymes. This has proved to be a rapid and sensitive technique for screening enzymes for enantioselectivity and for monitoring this throughout the course of the reaction.     

Immunochemical quantitation of dextran (α-1,6-glucan) traces in sucrose solutions by a rapid and sensitive automated nephelometric procedure

Dextran was quantitated in sucrose preparations using a polymer-enhanced immunonephelometric technique. The procedure is rapid, automated, and specific and has high sensitivity. The method shows excellent precision and good correlation with a reversed single radial immunodiffusion method. The dextran content of different saccharide preparations has been analyzed and the results are reported.