共查询到4条相似文献,搜索用时 2 毫秒
1.
Laura E. Edgington-Mitchell Nicholas Barlow Luigi Aurelio Aminath Samha Monika Szabo Bim Graham Nigel Bunnett 《Bioorganic & medicinal chemistry letters》2017,27(2):254-260
Activity-based probes are small molecules that covalently bind to the active site of a protease in an activity-dependent manner. We synthesized and characterized two fluorescent activity-based probes that target serine proteases with trypsin-like or elastase-like activity. We assessed the selectivity and potency of these probes against recombinant enzymes and demonstrated that while they are efficacious at labeling active proteases in complex protein mixtures in vitro, they are less valuable for in vivo studies. We used these probes to evaluate serine protease activity in two mouse models of acute inflammation, including pancreatitis and colitis. As anticipated, the activity of trypsin-like proteases was increased during pancreatitis. Levels of elastase-like proteases were low in pancreatic lysates and colonic luminal fluids, whether healthy or inflamed. Exogenously added recombinant neutrophil elastase was inhibited upon incubation with these samples, an effect that was augmented in inflamed samples compared to controls. These data suggest that endogenous inhibitors and elastase-degrading proteases are upregulated during inflammation. 相似文献
2.
Joana F. Lima Paulo Maia Beatriz T. Magalhães Laura Cerqueira Nuno F. Azevedo 《Biotechnology and bioengineering》2020,117(10):3212-3223
Fluorescence in situ hybridization (FISH) has been extensively used in the past decades for the detection and localization of microorganisms. However, a mechanistic approach of the whole FISH process is still missing, and the main limiting steps for the hybridization to occur remain unclear. In here, FISH is approached as a particular case of a diffusion-reaction kinetics, where molecular probes (MPs) move from the hybridization solution to the target RNA site within the cells. Based on literature models, the characteristic times taken by different MPs to diffuse across multiple cellular barriers, as well as the reaction time associated with the formation of the duplex molecular probe-RNA, were estimated. Structural and size differences at the membrane level of bacterial and animal cells were considered. For bacterial cells, the limiting step for diffusion is likely to be the peptidoglycan layer (characteristic time of 7.94 × 102 – 4.39 × 103 s), whereas for animal cells, the limiting step should be the diffusion of the probe through the bulk (1.8–5.0 s) followed by the diffusion through the lipid membrane (1 s). The information provided here may serve as a basis for a more rational development of FISH protocols in the future. 相似文献
3.
Philip E. Stanley 《Luminescence》1993,8(2):51-63
This survey was compiled between January and November 1992 from public domain information requested by the author from companies specializing in these products. It includes individual sections giving the name and address of each company together with brief details of their (a) kits used for assay of certain analytes in the routine laboratory and (b) reagents used mainly as research tools. This survey is a companion to a previous survey about 90 commercially available luminometers (Stanley PE. J Biolumin Chemilum 1992; 7: 77–108 and 157–69). 相似文献
4.
Jean Rivoal Christopher R Smith Trevor F Moraes David H Turpin William C Plaxton 《Analytical biochemistry》2002,300(1):94-99
We describe a method for the detection of isoforms of several glycolytic enzymes by activity staining after native PAGE. The staining is based on coupled enzyme assays carried out on the gel after electrophoresis and is linked to the disappearance of NADH, which is visualized by fluorescence. This method offers reliable and sensitive detection for phosphoenolpyruvate carboxylase, PPi-dependent phosphofructokinase, and pyruvate kinase from plant tissues. It can be applied to the detection of all enzymes which are normally detected spectrophotometrically using coupled enzyme assays consuming NAD(P)H. 相似文献