免疫学技术

In 1988 the first report on the expression of a fully functional recombinant Fab in E. coli (1) was one of the initiating events in the development of the field of antibody engineering. Shortly after came the report of the cloning and expression of antibody-variable domains as single-chain Fv or sFv (2). T ...

Introduction of mutations in antibody combining sites provides a powerful means to assess the functional role of individual residues and segments of the combining sites in interactions with antigen. Once cloned antibody fragments are available, several methods can be utilized to int ...

The display of antibody fragments on the surface of filamentous bacteriophages (1–7) constitutes a powerful system for the selection of molecules with desired specificities. In phage display, the antibody fragment is coupled to the minor coat protein (protein3) of bacteriophage M13 p ...

The application of phage display to the screening of antibody V-gene libraries promises to replace animal immunization and hybridoma technology as a means of antibody generation (1,2). Synthetic libraries offer the potential for expanding the immune system repertoire and designing ...

Asthma is associated with increased catalytic autoantibodies to vaso-active intestinal peptide (VIP), a bronchodilator and anti-inflammatory neuropeptide found in nerve endings supplying airway smooth muscle and epithelial structures (1). Asthmatic airways obtained at ...

Immobilized metal affinity chromatography (IMAC), introduced in 1975 (1), relies on the formation of coordinate bonds between metal ions immobilized on a suitable support and electron donor groups in proteins. A polyhistidine tag (five or six His residues) placed at either the C- or N-termin ...

A subpopulation of antibodies with high affinity for the substrate ground state is also capable of chemical catalysis (1). However, high turnover by catalysts is dependent on efficient binding to the transition state, not the ground state. Direct screening for catalysis may permit isolat ...

Polyclonal antibodies often serve as the starting point for interesting studies of new antibody functions and their links with immunoregulation and autoimmune disease. For example, the catalytic activity of naturally formed antibodies was first discovered using human autoant ...

High-specific-activity radiolabeled polypeptide probes permit sensitive detection of antigen-antibody interactions. In the case of small polypeptides labeled with 125I, various iodinated forms can be separated by reversed-phase HPLC and resolved peaks characterized by ...

Antibodies possess a vast repertoire of specificity and affinity. To understand the molecular basis of antibody function, we require high-resolution X-ray crystallographic structures and good solution structures of free and antigen-bound antibodies. The number of reported an ...

Radioimmunotherapeutic and radioimmunodiagnostic procedures have been the subjects of nearly 200 clinical trials since 1978. Applications range from diagnosis of tumor deposits to systemic radiation therapy. Originally, murine IgG was the most common antibody form used cli ...

A three-dimensional structure of reasonably high resolution is the starting point of any protein engineering project, whether intended to improve the affinity of an antibody, change the specificity of an enzyme, or to test a hypothesis regarding the role of a particular residue in determin ...

Antibodies are multisubunit proteins, but unlike many other multisubunit proteins, it is possible to study structural and functional properties of an antibody subunit free of the others. Thus, antibodies are singular proteins in that they are highly amenable to analysis by a fundamental ...

Antigen-binding sites in antibodies are formed by the variable regions of light (L) and heavy (H) chains. Delineation of antigen interactions with the individual subunits of antibodies is of interest for several reasons, including: 1. F

Major advances in the analysis of biomolecular structure have emerged from the development of highly specific monoclonal antibodies (MAb) by K�hler and Milstein in 1975 (1). MAb are produced from hybridoma cell lines that secrete a single species of antibody to a unique antigen. The advantag ...

This chapter was written with the express goal of introducing several basic concepts involved when antigens and antibodies interact in vitro to the novice reader and to serve as a review of these concepts to the more advanced reader. Most of these concepts also apply to in vivo reactivity. Antigen- ...

Bacterial endotoxin is probably the most common significant contaminant that might be found in antibody preparations. It is found ubiquitously in normal environments but its pyrogenic effects in vivo can be lethal. It is absolutely vital to control the level of endotoxin in therapeutic pr ...

Ideally, injectable drugs are sterilized in their final containers by a foolproof method like autoclaving. This is not possible for biologicals like monoclonal antibodies (mAbs), so they must be manufactured aseptically, sterilized by filtration and then filled into sterile vials or ...

During every stage of development and production of diagnostic or therapeutic antibodies, it is necessary to have an assay to measure antibody concentration. Several techniques are routine in virtually all antibody laboratories. High-performance liquid chromatograpy (HPLC) ...

Originally, digestion of antibodies by proteolytic enzymes was used to study their structure. Many diverse structures can be obtained by fragmentation of the different classes of antibody with different enzymes, or by using the same enzyme and changing the conditions Fig. 1). Not all the frag ...