Dextran matrix

PGIP, purified fi om P.vulgaris hypocotyls [11], was immobilized to the sensor ch via amine coupling. A continuous flow of HBS buffer (5 pl/min) was mantained over the sensor surface. The carboxylated dextran matrix of the sensor surface was first activated by a 6-min injection of a mixture of N-hydroxy-succinimide and N-ethyl-N - (3-diethylaminopropyl) carbodiimide, followed by a 7-min injection of PGIP (lOng/pl in 10 mM acetate, pH 5.0). Hie immobilization procedure was con leted by a 7-min injection of 1 M ethanolamine hydrochloride to block the remaining ester groups. 

Figure 5. The binding of antibodies to carboxymethyl dextran matrix attached to a sensor surface.

SPR chips coated with a carboxymethyl dextran matrix are supplied commercially by Biacore, a leading manufacturer of SPR instruments. Similar technique can be used for preparation of carboxymethyl dextran matrix on Me(Si)Ox surfaces on which -OH groups are generated by reaction with 3-glycidoxypropyltrimethoxysilane13. 

The enantiomorphs of a given sugar were found to exhibit no appreciable difference in Ka for example, the values for D- and L-mannose were 0.626 and 0.627, respectively. It appears, therefore, that stereospecific adsorption to the gel, which could possibly result from the presence of the dissymmetric residues of D-glucopyranose in the dextran matrix, is not a significant factor. 

In accordance with theoretical predictions of the dynamic properties of networks, the critical concentration of dextran appears to be independent of the molecular weight of the flexible polymeric diffusant although some differences are noted when the behaviour of the flexible polymers used is compared e.g. the critical dextran concentrations are lower for PEG than for PVP and PVA. For ternary polymer systems, as studied here, the requirement of a critical concentration that corresponds to the molecular dimensions of the dextran matrix is an experimental feature which appears to be critical for the onset of rapid polymer transport. It is noteworthy that an unambiguous experimental identification of a critical concentration associated with the transport of these types of polymers in solution in relation to the onset of polymer network formation has not been reported so far. Indeed, our studies on the diffusion of dextran in binary (polymer/solvent) systems demonstrated that both its mutual and intradiffusion coefficients vary continuously with increasing concentration 2. ... 

Crosslinking or entrapment Entrapment in dextran matrix, polyacrylamide or DNA crosslinked by using some agents such as glutar aldehyde [43] ... 

The mechanisms of drug release from dextran matrix occur in the early stage by polymer swelling, and the tablet thickness increases. Soon thereafter, polymer (and drug) dissolution starts occurring. The polymer dissolves because of chain disentanglement. Thus, there is a slow diminution of the thickness because of erosion until, finally, the tablet disappears (time > 480min). 

Studies of the interaction of IL-6 and the sIL-6R were monitored by SPR detection using a BIAcore instrument [1] (Pharmacia, Uppsala). Immobilisation of the respective proteins to the carboxymethylated dextran matrix coating the gold sensor chip was performed using the EDC/NHS coupling chemistry as previously described for IL-6 and sIL-6R [3, 7, 8]. Regeneration of the sensor surface for IL-6 and sIL-6R was performed with lOmM HCl for 3 min [8] and 4M MgCl2 in lOmM Tris-HCl buffer, pH 7.4 for 1 min respectively. 

All of the transport models presented so far assume that the diffusion mo-bihty of the analyte in the active sensor zone is the same as in the bulk. In case of the sensors using a thick skeleton to fix the receptors, such as a dextran matrix, solgel, or MIPS, it might be useful to take into account varying analyte diffusion mobility inside the active sensor layer. Detail analysis and proposed models can be foimd in [28]. 

The first SPR immunosensor for detection of pesticides was developed by Mimmni et al. [22] in the early 1990s. They used an SPR sensor developed by Biacore AB, Sweden, with the atrazine derivative bound to dextran matrix on the sensor chip. The detection of atrazine was performed using the inhibition assay and monoclonal antibodies. The sensor response was subsequently amplified by secondary antibody, which was bound to the antibody captured by the atrazine derivative (sandwich assay, see Chap. 7 in this volume [54]). This biosensor was demonstrated to measure atrazine in distilled and tap water within the range 0.05-1 ng mL in 15 min and exhibited relatively low crossreactivity with simazine and tetrabutyl atrazine (20%). The sensor surface was regenerated with 100 mM sodium hydroxide in 20% acetonitrile. 

Nakamura et al. demonstrated direct detection of herbicides by using a heavy-subunit-histidine-tagged photosynthesis reaction center (HHisRC) from bacterium Rhodobacter sphaerodies [26] (Fig. 3). They used a Biacore X instrument (from Biacore AB, Sweden) and a sensor chip with dextran matrix... 

Shimomura et al. used a Biacore 2000 SPR sensor instrument (Biacore AB, Sweden) for detection of PCB 3,3 4,4, 5-pentachlorobiphenyl [32]. They employed competition assay format and the sensor chip with polyclonal antibodies immobilized in the dextran matrix. The sample was mixed with a conjugate of PCB-horseradish peroxidase (HRP) and injected into the sensor. The presence of the analyte was detected as a decrease in binding of PCB-HRP conjugate. The detection was performed in 15 min with a detection limit of 2.5 ngmL in buffer. The sensor was demonstrated to be regenerable by 0.1 M hydrochloric acid. 

Detection of heavy metals was demonstrated by Wu et al. who used a Bi-acore X instrument (Biacore AB, Sweden) with rabbit metallothinein coupled to dextran matrix on the sensor chip [48]. Metallothinein is a protein that can be found in the cells of many organisms and is known to bind to metals (especially cadmium and zinc). Model experiments in which metallothein was used as a receptor demonstrated the potential of this sensor to directly detect Cd, Zn, and Ni in buffer at concentrations down to 0.1 jigmL . ... 

Isotype-specific anti-adenoviral antibodies in patients dosed with adenoviral-based gene therapy vectors were detected using a Biacore 3000 by Abad et al. [39]. In this assay, whole intact virus was immobihzed on the sensor surface (dextran matrix) using amine coupHng chemistry. The binding of antibodies from patient blood sera (1 10 diluted) was measured by SPR sensor and ELISA. The results obtained by the SPR biosensor were consistent with those obtained using ELISA. 

Figure 1 Strategies for GPCR attachment to dextran matrix. The interactive surface of a sensor chip consists of a self-assembled dextran monolayer (SAM) that bears functional groups to allow interaction of different compounds. For GPCR capture on the sensor chip surface, several strategies are used including direct binding to the dextran surface of monoclonal antibodies (A), avidin (B), GAG (C), or LVP (D).

Figure 8.27 Ds (left scale) of ( ) 864 kDa dextran probes through an aqueous 20.4 kDa dextran matrix (scale raised tenfold for clarity), and ( ) 71, (0) 148, and (O) 487 kDa dextrans through Myj 680 kDa hyaluronic acid, and Ds/Do ( , right scale) in 310 kDa polyvinylpyrrolidone of a 433 kDa dextran, using measurements by Daivis, et al.(38), De Smedt, et a/.(43), and Tinland and Borsali(44).

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