In this experiment, a novel biotin-avidin conjugation probe was synthesized and employed in the detection of reverse-phase protein microarray. Firstly, the proportion of the biotin-avidin conjugation probe was optimized. Then the rat IgG and goat anti-rat IgG system was served as a model to optimize the fabrication conditions of reverse-phase protein microarray, including the non-specific absor...

scAvd (single-chain avidin, where two dcAvd are joined in a single polypeptide chain), having four biotin-binding domains, was constructed by fusion of topologically modified avidin units. scAvd showed similar biotin binding and thermal stability properties as chicken avidin. The DNA construct encoding scAvd contains four circularly permuted avidin domains, plus short linkers connecting the fou...

The strong interaction between avidin and biotin is so tight (dissociation constant 10(-15) M) that conditions usually sufficient for protein denaturing fail to dislodge biotin from the avidin-biotin complex. This kind of irreversible binding hinders the use of avidin in applications such as affinity purification or protein immobilization. To address this concern, we have constructed a series o...

Affinity separation using functionalized microfiltration (MF) membranes provides a hydrodynamically favorable and cost-effective alternative to conventional column chromatography for the separation and purification of a specific protein from mixture of proteins. This type of separation in MF membranes is carried out by functionalizing the membrane with a suitable ligand containing specific inte...

To study the process of hormone action, we have developed an in vitro system utilizing minced oviduct from estrogen-treated chicks incubated in tissue culture medium. Progesterone added to the medium induced synthesis of a specific protein, avidin, that continued for up to 96 hr. During this period there was no increase in total oviduct protein, ovalbumin, or lysozyme, which suggests the specif...

Avidins are a family of proteins widely employed in biotechnology. We have previously shown that functional chimeric mutant proteins can be created from avidin and avidin-related protein 2 using a methodology combining random mutagenesis by recombination and selection by a tailored biopanning protocol (phage display). Here, we report the crystal structure of one of the previously selected and c...

Initial contact between the opportunistic fungal pathogen Candida albicans and host tissue occurs at the cell surface. Biotin derivatives have been used to label the cell-surface proteins of yeasts, with labelled proteins subsequently detected by avidin-reporter conjugates. Previous work has indicated that avidin can bind to C. albicans proteins in the absence of biotin, suggesting a possible h...

The paramagnetic microbead-based electrochemical binding assay was demonstrated for detecting two kinds of protein simultaneously. The principle of this assay is based on the sequestration electrochemistry. The protein binding electroactive magnetic microbeads which are conjugated with an electroactive compound and a ligand to bind specifically with a target protein were prepared. The avidin-bi...

We have developed an affinity biosensor system based on avidin-biotin interaction on a gold electrode. As the building block of an affinity-sensing monolayer, a fourth-generation (G4) poly(amidoamine) dendrimer having partial ferrocenyl-tethered surface groups was prepared and used. The unmodified surface amine groups from dendrimers were functionalized with biotinamidocaproate, and the biotiny...

Avidin is a tetrameric protein that belongs to the calycin superfamily. It has been studied mainly because of its extraordinary affinity to biotin, which led to a wide range of applications based on the avidin-biotin system. In the present study, we report the first crystal structures of avidin in a complex with two novel fluorescent pyrene derivatives: 1-biotinylpyrene (B9P) and 1-desthiobioti...