Biotinylated conjugate

Determine Vq and Vt by chromatography of 0.5 ml of Soln. A. The blue fractions represent that volume which will contain the biotinylated conjugate, and the red fractions will give the waste (unbound biotin). After chromatography, equilibrate the column again with PBS. 

Besides commercial kits, many immunochemical reagents are available (191,192) anti-drug antibodies, anti-enzyme antibodies, radioisotopic labels, nonisotopic labels, secondary antibodies, and biotinylated conjugates. Because of the flexibility in IA designs, investigators can consider several options from the availability of the commercial materials in addition to in-house resources in preparing the IA reagent components. 

The biotinylated conjugates (ACB, AC, BC, or C) are captured by a sensor chip, whose surface is coated with immobilized strept-avidin and which acts via surface plasmon resonance as a tool for enzyme (R) binding experiments. 

P-elimination, 266 benzyl ester, 267, 274 biotinyl conjugate, 270 chemoselectivity, 268 compatibility, 268 enz3unatic reactions, 267, 268 ethyl 2-ethoxy-1,2-dihydroqoinoline-1-caiboxylate (EEDQ), 271 9-fluorenylmethoxycarbonyl (Fmoc) group, 266, 267, 268, 269, 273, 274, 276... 

Lacto-iV-neotetraose with X (see above) = P-D-GICA3-SO3H has been made as a 2-aminoethyl glycoside and converted to biotinylated conjugates. ... 

Then they reported an efficient and stereocontrolled construction of homo- and heterogeneously 4-and 6-sulphated CS oligomers as their biotinylated conjugates from suitable protected disaccharide building blocks. A series of sulphated and unsulphated analogues of the linkage oligosaccharide and of the constitutive unit of CS has been synthesized. ... 

Avidin (occurring in eggs) or streptavidin (produced by Streptomyces avidinii) are usually tetrameric proteins, where each subunit has very high affinity to bind biotin (vitamin B7). The formation constants lie between 10 " and 10 which can be classified as the strongest noncovalent bonds. Thus, biotinylated conjugates can be easily bound to avidin-modified counter-partners and surfaces. 

An enzyme-amplified detection scheme, based on tire coupling of a streptavidin-alkaline phosphatase conjugate and biotinylated target sequences was then applied. The enzyme catalysed the hydrolysis of the elecn oiiractive a-naphthyl phosphate to a-naphtlrol this product is elecU oactive and has been detected by means of differential... 

FIG. 6 Successive coupling of two different biotinylated compounds with the DNA-STV conjugates 2 [33]. In a first step, a macromolecular functional component (FC, represented by the shaded ellipse), such as a biotinylated enzyme or oligonucleotide, is coupled. In a second step, a biotinylated low-molecular-weight modulator M, represented by the shaded sphere) is coupled to the remaining free biotin-binding sites. The modulator is used to modify the conjugate s hybridization properties or to supplement its functionality. 

FIG. 8 Synthesis of oligomeric DNA-STV conjugates 7 from 5, 5 -bis-biotinylated DNA and STV [49]. Note that the schematic structure of 7 is simplified, since a portion of the STV molecules function as tri- and tetravalent linker molecules between adjacent DNA fragments. 

Streptavidin-single-stranded DNA covalent conjugates were described as the building blocks for assembling nanostructured scaffolds [31], The amount and type of biotinylated ligands were used to modulate the affinity of duplex formation between solid-phase-bound nucleic acid templates and DNA-streptavidin conjugates. This system has been proposed for the design of fine-tuned sequence detection systems. 

Patterning of enzyme monolayers on a solid surface was carried out by photoactivation of immobilized monolayer of caged -biotin derivatives in selected areas. Specific oriented binding of enzyme-avidin conjugates could be readily made to the photoactivated zones [42]. Oriented immobilization of G-protein-coupled receptors on a solid surface was also made possible on a biotinylated surface by first immobilizing streptavidin, followed by the immobilization of biotinylated G-protein-coupled receptor [43]. 

The LANCE cAMP assay is a competitive assay in which cAMP produced by the cells competes with fluorescent-labeled acceptor cAMP for a cryptate tagged donor antibody. The principal of the assay is shown in Fig. 6. On the left strepta-vidin conjugated Europium binds to biotinylated cAMP. An antibody labeled with the fluorescent dye Alexa binds to the cAMP, bringing the donor and acceptor into close proximity, and energy transfer occurs. When the cell releases cAMP, it competes with the biotin-labeled cAMP for the antibody, and a signal decrease is observed. In the TR-FRET assay the antibody is directly labeled with either Eu or Tb. In this format an increase in cAMP also causes a decrease in signal. 

A similar type of biotin-dendritic multimer also was used to boost sensitivity in DNA microarray detection by 100-fold over that obtainable using traditional avidin-biotin reagent systems (Stears, 2000 Striebel et al., 2004). With this system, a polyvalent biotin dendrimer is able to bind many labeled avidin or streptavidin molecules, which may carry enzymes or fluorescent probes for assay detection. In addition, if the biotinylated dendrimer and the streptavidin detection agent is added at the same time, then at the site of a captured analyte, the biotin-dendrimer conjugates can form huge multi-dendrimer complexes wherein avidin or streptavidin detection reagents bridge between more than one dendrimer. Thus, the use of multivalent biotin-dendrimers can become universal enhancers of DNA hybridization assays or immunoassay procedures. 

Biotinylated dendrimers also have been used to develop targeting conjugates for therapeutic use in targeting cancer cells. Wilbur et al. (1998) studied several different PAMAM... 

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