PRODUCTION OF SINGLE-CHAIN ANTIBODIES AGAINST HUMAN FERRITIN IN ESCHERICHIA-COLI

Hybridoma cells capable of producing antibodies against human ferritin were used for cloning cDNA copies of immunoglobulin heavy and light c hain genes. By means of the polymerase chain reaction (PCR) we obtaine d the variable segments of these genes. Specific amplifications of the ligase reaction products were performed to combine the variable segme nts with the DNA fragments encoding the peptide linker and the signal peptide of the cloned Erwinia carotovora pelB gene. Upon expression of the gene encoding the single-chain antiferritin antibodies under cont rol of T7 RNA polymerase in Escherichia coli, the processed molecules of recombinant proteins formed aggregates in the periplasm of producin g cells. Reversible denaturation in the absence of reducing agents all owed us to obtain single-chain antibodies with the original binding sp ecificity to human ferritin.