Stopped-flow system

Depending on the configuration of the device and the method for sample and reagents introduction, it is possible to classify the systems into static (batch or discrete sampling instrument) or flowing stream, both using continuous-flow or stopped-flow systems. 

The quantum yield of the mant fluorophore in mant-GDP or mant-GTP increases approximately by 100 % when the molecule changes from the aqueous environment into the nucleotide binding pocket of the Ras protein. Therefore the kinetics of complex formation between nucleotide free Ras and the mant analogues of GDP or GTP could be detected easily in a stopped flow system by an increase in fluorescence signal. 

The possibility for automation is offered by the Voyager SF with its stop flow system, able to process larger reaction volumes by sequen-... 

A number of stopped-flow systems are commercially available. Three of the most used are manufactured by Atago Bussan (formerly Union Giken), Japan Dionex (formerly Durrum), USA and Hi-Tech Scientific, UK. These also manufacture rapid scan spectrophotometers, multimixer, temperature-jump and flash photolysis equipment. 

Figure 2.15 — Variants of coupled continuous configurations and sensors accommodating an immobilized catalyst (Cat). (A) Conventional system. (B) Stopped-flow system. (C) Configuration with iterative reversal of the flow direction. (D) Open-closed circuit configuration. Symbol meanings are given in Fig. 2.14. For details, see text.

B. Serra, A.J. Reviejo, C. Parrado and J.M. Pingarron, Graphite-Teflon composite bienzyme electrodes for the determination of L-lactate application to food samples, Biosens. Bioelectron., 14(5) (1999) 505-513. A.A.J. Torriero, E. Salinas, F. Battaglini and J. Raba, Milk lactate determination with a rotating bioreactor based on an electron transfer mediated by osmium complexes incorporating a continuous-flow/ stopped-flow system, Anal. Chim. Acta, 498(1-2) (2003) 155-163. 

A.A.J. Torriero, E. Salinas, F. Battaglini and J. Raba, Milk lactate determination with a rotating bioreactor based on an electron transfer mediated by osmium complexes incorporating a continuous-flow/stopped-flow system, Anal. Chim. Acta, 498 (2003) 155-163. 

Fig. 3.7 Generic layout of a single-mixing stopped-flow system.

Before any instrument can be used with confidence its operating characteristics must be well understood. The exceptional versatility of the vidicon rapid scanning stopped-flow system allowed a large variety of characterization experiments to be easily performed. A few of the more important experiments designed to help characterize the vidicon detector will be presented before the actual application of the instrument is discussed. 

Figure 3.7. Commercial stopped-flow system for fast liquid-phase reactions (courtesy KinTek Corporation, reproduced with permission).

Electronic absorption spectroscopy can be used to determine the general structure of porphyins and their derivatives. The oxidation and coordination state of the iron and the identity of the amino acid that ligates the heme can be examined by comparing the absorption spectrum of the protein of interest with the spectra of known heme proteins (45, 65). General characterization with electronic absorption spectroscopy indicated that NO and CO, but not O2, bind to the sGC heme moiety (66). Dynamic studies of ligand binding and dissociation also can be examined with this technique on psec-msec time scales with standard stopped-flow systems. 

The same electrochemical flow cell has also been used in combination with IR spectroscopy in a stopped-flow system [68, 69]. The purpose was to study the electrogeneration of the Collman reagent Fe(CO)4 obtained from the two-electron reduction of Fe(CO)5. It was possible to monitor the consumption of Fe(CO)s and... 

Figure 9. Thermal stopped-flow system (a) conventional stopped-flow (b) zero pressure drop stopped-flow.

Rapid-Mixing, Rapid-Scanning Stopped-Flow Systems... 

Rapid-Mixing, Rapid-Scanning Stopped-Flow Systems 3.3.1. SINGLE ELEMENT DETECTOR SYSTEMS... 

Inherently, the FI A stopped-flow procedure should be an ideal vehicle to determine reaction rates and rate laws, provided that an experimental approach could be designed that allows resolving the individual contributions of physical dispersion and chemical kinetics. A comprehensive treatment of this problem was recently described by Hungerford et al. [838], who pointed out that although the single-line stopped-flow system (Fig. 4.15a cf. Fig. 4.11) allows optimization of solution conditions for measurement during a selected stopped-flow time interval (fs) by choosing... 

When we examine the time distribution of the recorded signal (Fig. 4.18c), it is apparent that the peak width at its base in this system is approximately twice as long as the time interval between the sample injection and the moment when the signal starts to rise. In a simple stopped-flow system this time distribution does not present a problem because a new cycle is initiated only when the next sample is injected. The timer for a two-channel parallel stop analysis must, however, run in a fixed go-stop-go cycle, because while one sample zone is being injected (or flushed out) the other is stored, and if the length of the storage period is not always the same, the analyzer will not yield reproducible results-unless, of... 

A stopped-flow setup can relatively easily be equipped with multispectral detection. The fluorescence light is emitted from a small spot in the flow channel. Therefore the flow cell can be placed directly in the input slit plane of a poly-chromator. The spectram is detected by a multianode PMT. The photons detected in the spectral channels are recorded simultaneously by a TCSPC device and a router. However, although the implementation is relatively simple, no spectrally resolved TCSPC-based stopped-flow system has yet been described. 

FIGURE 3.4 A schematic drawing of a stopped flow system with the source and detector of the spectrophotometer indicated. 

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