Single-chain urokinase plasminogen activator (scu-PA) that had been modified with N-succinimidyl-3-(2-pyridyldithio)propionate was covalently linked by disulfide bonds to the Fab' of a monoclonal antibody specific for the beta-chain of fibrin (antibody 59D8). scu-PA-59D8 Fab' conjugate was separated from free scu-PA and two-chain urokinase coupled to 59D8 Fab' by two-step affinity chromatograph...

Active site structure-function analyses of anti-fluorescein single chain antibody 4-4-20 and anti-single-stranded DNA single chain antibody 04-01 were conducted studying the ligand binding properties of hybrid antibodies resulting from systematic transplantation of heavy chain complementarity regions (HCDRs) from monoclonal antibody 4-4-20 into 04-01. Two prototype monoclonal antibodies were ch...

The objective of this study was to identify and construct a human natural phage single-chain antibody (scFv) library of human anaplastic thyroid carcinoma (ATC) using phage display technology. Total RNA was extracted from lymphatic tissue near an ATC and used to amplify variable heavy chain (VH) and variable light chain (VL) fragments with added linker sequences using reverse transcription-poly...

Single-chain Fv (sFv) proteins are genetically engineered molecules that consist of the two variable domains of an antibody connected by a polypeptide linker; they contain the antigen binding function of the parental protein in a single 30-kDa polypeptide chain. sFvs are usually produced in bacteria where they are insoluble and therefore require extensive refolding in vitro. In this report we f...

Three foldases--the apical domain of GroEL (mini-chaperone) and two oxidoreductases (DsbA and DsbC) from Escherichia coli--were studied in refolding a protein with immunoglobulin fold (immunoglobulin-folded protein) that had been produced as inclusion bodies in E.coli. The foldases promoted the refolding of single-chain antibody fragments from denaturant-solubilized and reduced inclusion bodies...

There is emerging evidence that the misfolding of superoxide dismutase 1 (SOD1) may represent a common pathogenic event in both familial and sporadic amyotrophic lateral sclerosis (ALS). To reduce the burden of misfolded SOD1 species in the nervous system, we have tested a novel therapeutic approach based on adeno-associated virus (AAV)-mediated tonic expression of a DNA construct encoding a se...

A murine hybridoma cell line has been established that consistently forms large cytoplasmic inclusions. These structures bind antibody specific for mouse kappa L chain when stained in situ. SDS-PAGE analysis of isolated inclusion bodies produce a single protein band of approximately 26,000 Mr that reacts with anti-kappa antibody when transferred to nitrocellulose. No carbohydrate was detected i...