Preparation of temperature-sensitive antibody fragments

We designed a temperature-sensitive single chain, antibody fragment in which the antigen-binding activity was drastically altered by a temperature-shift between 4 and 37°C. An oligonucleotide corresponding to the temperature-sensitive helix-coil transition peptide,-(-Glu Ala Ala Ala Lys-)-, was introduced between Vh and Vl genes of scFv against the simian virus 40 large T antigen. The antigen-binding activity of the temperature-sensitive scFv produced by Escherichia coli cells showed a maximum decrease of 1/36 at 4°C compared with that at 37°C. Binding activity was controlled by the NaCl concentration, as well as by the temperature shift. By using a column that immobilized the ScFv, the antigen was purified in response to elution by the temperature-shift.