细胞技术

monolithic macroporous hydrogels, “cryogels,” are produced by polymerization in a partially frozen state when the ice crystals perform as a porogen. Cryogels have a unique combination of properties: (i) large (10–100 μm) pores; (ii) minimal non-specific interactions due to the hydroph ...

Methods are presented for the preparation, ligand density analysis and use of an affinity adsorbent for the purification of a glutathione S-transferase (GST) fusion protein in packed and expanded bed chromatographic processes. The protein is composed of GST fused to a zinc finger transcr ...

Enzymes immobilized on solid-phase matrices have found various applications in biotechnology, molecular biology and molecular diagnostics and can serve as industrial catalysts and as specific reagents for analytical procedures. A wide range of supports have been utilized for i ...

The interaction between avidin and biotin or streptavidin and biotin forms the basis of several widely used immunohistochemical techniques. An assumption inherent to these techniques is that endogenous biotin is not present in the tissue in detectable quantities, as neither avidin n ...

Immunodetection techniques are dependent on enzyme–protein conjugates for the visualization of antigen–antibody complexes. One of the most widely used is the avidin–biotin–peroxidase complex (ABC) method. However, treatment of certain tissues with ABC reagents alone may res ...

Posttranslational modifications of histones play an important role in the regulation of chromatin structure and, hence, gene regulation. Recently, we have identified a novel modification of histones: binding of the vitamin biotin to lysine residues in histones H2A, H3, and H4. Here, we des ...

The biotin–avidin microplate assay is a sensitive method to measure methylation of biotinylated oligonucleotide substrates by DNA methyltransferases (MTases). The methylation reaction is carried out in solution using -AdoMet. Afterwards, the oligonucleotides are immob ...

A number of endogenously biotinylated proteins are found in both cytosol and mitochondria of mammalian cells from many tissues, including liver, spleen, pancreas, kidney, and intestine. Therefore, caution should be taken when using the biotin detection system. Endogenous biotin can i ...

Carbohydrates represent a fundamental building unit of living organisms. Many contemporary results introduce these substances as medium with remarkable data storage capacity (glycocode) that is decoded by special receptor counterpartners, lectins. Animal so-called end ...

Biotinylation of proteins is a powerful tool for investigating biological phenomenon, both in vitro and in vivo. Biotinylating reagents that form covalent bonds with several types of amino acid residues are commercially available. However, most, if not all, of these commercially avail ...

Analytical methods characterizing the immunogenicity of antigens are useful for monitoring, characterizing and predicting antibody responses to therapeutic biologics or vaccines. Distinct Luminex� microspheres coupled with protein G, anti-human immunoglobulin (Ig ...

Biospecific interactions are used in many capturing and bioseparation steps. A typical situation is the coupling of a biospecific ligand to a chromatographic stationary phase for affinity chromatography. This approach has two possible drawbacks. The first is that a chromatograph ...

Numerous methods have been published for the detection of protein using avidin–biotin technology. Complications can arise using this system when the protein of interest is in extremely high or low abundance. The ability to successfully detect high- or low-abundance proteins is depend ...

Avidin functional affinity electrophoresis (AFAEP) is a variational method of affinity electrophoresis. In this technique, avidin is immobilized within a small area of the gel matrix by interaction with acrylamide and/or polyacrylamide either directly or through bifunctional ...

Site-directed mutagenesis or directed evolution of proteins often leads to the production of inactive mutants. For streptavidin and related proteins, mutations may lead to the loss of their biotin-binding properties. With high-throughput screening methodologies in mind, it is im ...

Biotin-4-fluorescein (B4F) is a convenient molecular probe for (strept)avidin and for unlabeled biotin in homogeneous fluorescence assays. The primary standard is a 16 μM working solution of d-biotin which is used to titrate an aliquot of a (strept)avidin stock solution while monitori ...

The strong interaction between streptavidin and biotin is one of the most commonly exploited tools in chemistry and biology. Methods for the facile derivatization of a variety of molecules (in particular, proteins) with biotin have been introduced, in order to allow their efficient recov ...

The overexpression of toxic recombinant proteins is often problematic, leading to either low production levels or inclusion bodies. Streptavidin is no exception and thus the highest production level reported to date for streptavidin is 70 mg/L of functional protein. Herein, we report on ...

This chapter introduces the problem of ancestral sequence reconstruction: given a set of extant orthologous DNA genomic sequences (or even whole-genomes), together with a phylogenetic tree relating these sequences, predict the DNA sequence of all ancestral species in the tree. Blanc ...

Mitochondrial genomes (mtDNA) of animals are circular molecules of relatively small size, compactly organized, and generally encoding genes for 2 rRNAs, 22 tRNAs, and 13 proteins that are required for mitochondrial function. Methods of mtDNA isolation take advantage of its physical lo ...