Immobilization carbodiimide activation

On the other hand, covalent agents can also be applied to the unpreated carbon surface directly before DNA immobilization onto activated sites of carbodiimide compounds [21]. 

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 8 Chemiluminescent (A and B) and bioluminescent (C) detections for immobilized hybridizations of PCR product. Dg, digoxigenin Bt, biotin Ad, avidin. Procedure A [30] Biotin moiety is incorporated into PCR products during the amplification reaction, using one 5 -biotinylated primer. The product is hybridized with a Dg-labeled probe and is immobilized on streptavidin-coated magnetic beads. This capture reaction is carried out for 30 min at 37°C. A permanent magnet is used to sediment the beads during washing to remove unbound DNA. By incubation with the washed beads for 45 min at 37°C, anti-Dg antibody conjugated to HRP enzyme is bound to the Dg-labeled probe, and luminol reaction is performed for CL detection. Procedure B [31] Wells of apolystyrene microtiter plate are activated with l-ethyl-3-(3-dimethylaminopropyl)-carbodiimide, and then coated with a labeled cDNA probe complementary to an internal region of the target DNA.

Hirano et al. [150, 151] immobilized several peptides, RGDS, on ethylene-acrylic acid copolymer (EAA, acrylic acid content 20 wt%) film by reacting the amino-terminal of the peptide with the carboxylic acid of the copolymer with the aid of a water-soluble carbodiimide, to form EAA-co-NH-RGDX. Their objective was to examine effect of the fourth residue, X, on the cell-attachment activity of the tetrapeptide, RGDX, where X is S, V and T. They also examined the activity of RGD, YIGSR and YIGSR-NH2 for comparison. The cell lines used were ovary CHO-K1 cell (Chinese hamster), kidney NRK cell... 

A simple, one-step immobilization technique is bulk cross-linking of the functional protein with bovine serum albumin (BSA) using glutaraldehyde as the cross-linking agent (Fig. 2.19). It is popular because of its simplicity, but it usually leads to reduction of the biological activity of the biomolecule. A cleaner and preferable approach is the two-step carbodiimide route. A partial summary of the various immobilization options was shown in Table 2.3. 

Following the corona-discharge treatment of a silicone surface and the subsequent graft polymerization of AAc, type I atelocollagen was immobilized onto the grafted surface with the use of water-soluble carbodiimide [176, 177]. As depicted in Fig. 17, the immobilization reaction involves two steps, i.e., activation of carboxylic acids and the following nucleophilic substitution with prima-... 

Several useful schemes for attaching nucleic acid probes onto electrode surfaces have thus been developed [2-8]. The exact immobilization protocol often depends on the electrode material used for signal transduction. Common probe immobilization schemes include attachment of biotin-functionalized probes to avidin-coated surfaces [15], self-assembly of organized monolayers of thiol-functionalized probes onto gold transducers [16], carbodiimide covalent binding to an activated surface [17], as well as adsorptive accumulation onto carbon-paste or thick-film carbon electrodes [15-30]. 

Fig. 4 Nucleic acid immobilization. Immobilization of 5 -amino-C6 modified probes onto (a) carboxylate modified latex beads (1 im) via carbodiimide reaction and (b) Cyanogen bromide (CNBr) activated Sepharose beads (100 un). (0 cyclic structure, e.g. dicyclo-hexylcarbodiimide)...

Covalent immobilization of enzymes increases their stability while lowering their activity. Also, their storage stability is notably higher. The synthesis of arginine from citrulline, ATP and argenino-succinate synthetase may involve a carbodiimide intermediate. ... 

---