抗体应用

Functional characterization of antibodies that inhibit soluble cytokines or chemokines requires robust, sensitive in vitro and in vivo bioassays. Testing an antibody in vitro requires consideration of antigen source, integrity, and concentration, as well as the magnitude of the b ...

Neutralization tests are performed in vitro, at the cellular level - including on cells in the form of organs - and at the sub-cellular level. They assess the efficacy of antibody fragments, or antibodies, to inhibit critical stages of activities. These activities are generally deleterious to H ...

The use of radioiodinated proteins is one of the most sensitive methods to demonstrate and quantify specific protein/protein interactions. The direct introduction of Iodine-125 into proteins is easily and quickly done, provided the mandatory safety equipment is available. Most an ...

To determine the affinity, the KD,-value, between interaction molecules a number of techniques using labels are available, such as chromatography, isothermal titration calorimetry, radioimmuno- assay and the widely used enzyme-linked immunosorbent assay (ELISA). However, ne ...

The hexahistidine tag is one of most commonly used fusion tags in affinity purification of recombinantly expressed proteins. Real-time binding analysis using Biacore technology allows in-depth characterization of respective association and dissociation patterns of pote ...

Antibodies and their fragments are widely used tools for research, diagnostics and therapy. The strength of the interaction between an antibody molecule and the recognized antigen is conveniently characterized by the affinity constant. Among the available approaches to determi ...

Glycosylation is one of the main IgG post-translational modifications and has essential roles in antibody effectors functions, immunogenicity and plasmatic clearance. In this chapter we discuss and provide detailed protocols for IgG homogeneity and determination of level of gl ...

Among all analytical methods used to characterize monoclonal antibodies (mAbs), mass spectrometry plays an increasingly important role for both global and fine structural characterization of therapeutic candidates. In this chapter we discuss and provide detailed protocols ...

Size exclusion chromatography is a standard chromatographic technique that allows the separation of molecules or molecule complexes by their hydrodynamic volume (grossly equivalent to the molecular mass). In antibody engineering, it is used to analyze and to separate antibodies ...

Many approaches for antibody epitope mapping utilize peptides or peptide fragments and are limited to antibodies binding linear epitopes or epitopes not requiring fully folded conformationally correct antigens. The protocols in this chapter show how yeast displayed antigens a ...

Affordable high-density peptide arrays are needed to routinely define the exact binding sites of antibodies. In terms of prize and density peptide arrays currently lag far behind oligonucleotide arrays that are available in densities exceeding 50.000 oligonucleotides per cm2. This ...

Antibodies recognize and bind their target protein antigens via surface accessible interaction sites, the epitopes (Fig. 35.1), which can involve amino acid side chain and backbone contacts along a linear segment of the antigen amino acid chain (linear epitopes) or which can involve amino a ...

The following chapter describes the purification of whole IgG by Protein A affinity chromatography from crude protein mixtures such as serum, ascites fluids or cell culture supernatant. This reliable method is based on the ability of Protein A to bind with high affinity to the Fc region of immuno ...

Production of IgG antibodies is usually done in mamalian cells. Predominantly, chinese hamster ovary (CHO) cells are used for generating stably transfected cell lines and for manufacturing. In addition to large scale manufacturing, stably transfected CHO cells are also well suited for ...

Antibodies have come to the forefront of protein therapeutics in the past decade and while they are now widely accepted and administered to patients, the cost of using antibody therapy remains extremely high. Here we describe a protocol for the expression of human antibodies using the chlorop ...

Plants are attractive vehicles for the expression of recombinant proteins as they are inexpensive and versatile systems, amenable to rapid and economic scale-up. Furthermore, transgenic plants can be established in resource-poor areas, where the demand for pharmaceutical prote ...

Although natural IgA antibodies constitute an important immune defense mechanism at serosal surfaces, their immunotherapeutic potential has not been thoroughly explored. Among the limitations hampering the development of therapeutic IgA antibodies is the lack of well estab ...

Despite antibody fragments like scFv or Fab are sufficient for various tasks, complete recombinant IgG is sometimes needed. Production of IgG molecules is not generally achievable in E. coli. Insect cells are able to produce IgG and the use of the Baculovirus systems allows a rapid cloning and pro ...

The development of antibody-directed enzyme prodrug therapy is reviewed. Antibody-directed enzyme prodrug therapy (ADEPT) generates a cytotoxic drug selectively within deposits of cancer and is designed to give therapy with high levels of tumour selectivity coupled with excep ...

Interaction of antibodies with the neonatal FcRn receptor controls largely the length of their life cycle. Altering the serum persistence of antibodies through modulation of their interaction with the FcRn may be advantageous for achieving high contrast images shortly after appli ...