摘要:

The proven ability of insect cells to produce murine and human antibodies renders the baculovirus system amenable to the synthesis of both naturally occurring antibodies and designed mutants. Study of these antibodies should provide a basis for rational antibody design useful in human therapy.

ISSN:0883-0185    

DOI:10.3109/08830189309061688

出版商:Taylor&Francis    

年代:1993

数据来源: Taylor

摘要:

We review here our attempts to achieve a better understanding of the structure—function relationship of antibody combining sites, and to gain insights into the engineering of antibodies with desired specificity and affinity. We have focused on a model system—antibodies to the hapten p-azophenylarsonate (Ars) derived from A/J mice. Oligonucleotide-directed mutagenesis was used to alter the sequence of the variable region genes of such anti-Ars antibodies. Mutant antibodies were generated in hybridoma cells following transfection of the altered genes, and the effects of the primary structure changes on antibody specificity, affinity, and idiotypic expression were assessed. These studies suggest that an antibody combining site with basic specificity for an antigen could be created by introducing a set of a few amino acid residues in the complementarity determining regions, and that the affinity of such a site could be improved one substitution at a time in a sequential manner.

ISSN:0883-0185    

DOI:10.3109/08830189309061689

出版商:Taylor&Francis    

年代:1993

数据来源: Taylor

ISSN:0883-0185    

DOI:10.3109/08830189309061690

出版商:Taylor&Francis    

年代:1993

数据来源: Taylor

ISSN:0883-0185    

DOI:10.3109/08830189309061691

出版商:Taylor&Francis    

年代:1993

数据来源: Taylor

摘要:

Combinatorial antibody libraries, in which PCR amplified immunoglobulin light and heavy chain DNA are randomly recombined irrespective of their pairing in vivo into a vector and subsequently expressed in E. coli, have quickly become a very productive tool to generate monoclonal antibodies from various species. It has been drastically improved by utilizing phage display technologies in the selection process of specific antibodies. A brief summary of current techniques, critical published experiments showing the versatility of these systems with emphasis on human antibodies and discussions on chain preference, affinity maturation and the advent of semisynthetic and non-immune libraries will be presented.

ISSN:0883-0185    

DOI:10.3109/08830189309061692

出版商:Taylor&Francis    

年代:1993

数据来源: Taylor

ISSN:0883-0185    

DOI:10.3109/08830189309061693

出版商:Taylor&Francis    

年代:1993

数据来源: Taylor

摘要:

of the major advantages of genetic engineering is the ability to produce novel, hybrid antibodies. Hybrid antibodies can be assembled using fragments from different antibodies with the objective of assembling novel combinations of antibody-related effector functions. To efficiently achieve this goal it is necessary to have a precise understanding of the structure-function relationships within the antibody molecule. Secondly, it is possible to produce hybrids of antibodies with non-immunoglobulin proteins thereby achieving unique combination of functional properties. In this case it is necessary to consider both the desired functional properties and the means of assembling the protein components so as to maintain these properties. In all cases it is necessary to have the cloned gene segments, appropriate vectors and expression systems.

ISSN:0883-0185    

DOI:10.3109/08830189309061694

出版商:Taylor&Francis    

年代:1993

数据来源: Taylor

ISSN:0883-0185    

DOI:10.3109/08830189309061695

出版商:Taylor&Francis    

年代:1993

数据来源: Taylor

摘要:

A single-chain antibody or single-chain Fv (sFv) incorporates the complete antibody binding site in a single polypeptide chain of minimal size, with an approximate molecular weight of 26,000. In antibodies, the antigen combining site is part of the Fv region, which is composed of the VHand VLvariable domains on separate heavy and light chains. Efforts over nearly two decades have indicated that Fv fragments can only rarely be prepared from IgG and IgA antibodies by proteolytic dissection. Beginning in 1988, single-chain analogues of Fv fragments and their fusion proteins have been reliably generated by antibody engineering methods. The first step involves obtaining the genes encoding VHand VLdomains with desired binding properties; these V genes may be isolated from a specific hybridoma cell line, selected from a combinatorial V-gene library, or made by V gene synthesis. The single-chain Fv is formed by connecting the component V genes with an oligonucleotide that encodes an appropriately designed linker peptide, such as (Gly4-Ser)3. The linker bridges the C-terminus of the first V region and N-terminus of the second, ordered as either VH-linker-VLor VL-linker-VH. In principle, the sFv binding site can faithfully replicate both the affinity and specificity of its parent antibody combining site, as demonstrated in our model studies with the 26-10 anti-digoxin sFv. Furthermore, the sFv remains stable at low concentrations that promote VHand VLdissociation from the Fv heterodimer, resulting in loss of Fv binding. Intravenously administered sFv proteins exhibit accelerated biodistribution and exceptionally fast clearance compared to IgG or Fab. These pharmacokinetic properties allow rapid imaging by sFv, which therefore may be labeled with a shortlived isotope such as Tc-99m. Expression of a single gene product from fused sFv and effector genes facilitates immunotargeting of the effector protein, as shown for single-chain Fv toxin fusion proteins.

ISSN:0883-0185    

DOI:10.3109/08830189309061696

出版商:Taylor&Francis    

年代:1993

数据来源: Taylor

ISSN:0883-0185    

DOI:10.3109/08830189309061697

出版商:Taylor&Francis    

年代:1993

数据来源: Taylor