WebJun 1, 2002 · The in vivo assay uses CCF2/AM (E) (adapted from ref. 6). When no β-lactamase activity is present, excitation of the coumarin at 409 nm leads to FRET to the fluorescein acceptor and emission of ... WebSep 13, 2007 · The fluorometric assay is performed with a cephalosporin derivative, CCF2/AM, as the substrate . CCF2/AM contains ester moieties, which facilitate its …

β-Lactamase protein fragment complementation assays as

Webanalysis protocol. Widely used to assay injection of effector proteins in T3SS and T4SS is an assay that utilizes the enzymatic cleavage of the FRET-reporter cephalosporin CCF2 by injected β-lactamase fu-sions (Charpentier & Oswald, 2004). The CCF2 assay facilitated the analysis of injection kinetics and of intracellular accumulation levels WebDec 15, 2008 · CCF2 fluorescence readings were normalized by dividing fluorescence values measured at 447 nm by those measured at 520 nm, and then normalized to OD 600 of the cells. All data was then normalized to the background fluorescence measured in cells expressing cutinase only. flyway springboot 配置

A NanoLuc luciferase-based assay enabling the real …

WebThe CCF2 assay facilitated the analysis of injection 95kinetics and of intracellular accumulation levels of effectors (26). It was also successfully used for HTP high content screening of T3SS inhibitors96 (27). Real-time observation of injection was achieved by direct fluorescent labeling of tetracysteine motif-tagged effectors97 (28). However, WebThis cleavage changes the fluorescence emission spectrum of CCF2-AM from green (520 nm) to blue (447 nm), and thus permits detection of fusion by fluorescence microscopy, … WebNov 13, 2009 · This assay also serves as an orthogonal counterscreen to identify epi-absorbance assay artifacts. This biochemical FRET-based assay employs as the beta-lactamase substrate the cephalosporin … flyways software and solutions