Expression of a single-chain Fv antibody fragment specific for the hepatitis B surface antigen in transgenic tobacco plants

An anti-Hepatitis B virus surface antigen (HBsAg) single chain Fv (scFv) antibody fragment was expressed in Nicotiana tabacum transgenic plants. The 6-histidine tagged scFv was targeted to either the cytosol, apoplast, and vacuole, or for retention in the endoplasmic reticulum. Expression of active scFv was detected by ELISA in fresh leaf material from F1 transgenic plant lines representative of the genetic constructs targeting the antibody fragment to the apoplastic fluid (AF-12, 0.031% of the total soluble protein), vacuole (V-20, 0.032% of the total soluble protein), and endoplasmic reticulum (ER-52, 0.22% of the total soluble protein). No scFv was detected by ELISA or western blot in the plants transformed with the cytosol construct. The biologically active scFv was easily purified (to 94-95% purity) from ER-52 and AF-12 plant material using immobilized metal ion affinity chromatography. Recovery estimated from the ER-52 plant line indicates that 15-20 μg of pure active scFv can be obtained per gram of fresh leaf material, on a laboratory scale.