Soluble expression and target study to brain of anti-TfR ScFv.
- Author:
Bing YAN
1
;
Heng-Qi ZHU
;
Pei-Tang HUANG
Author Information
1. Beijing Institute of Biotechnology, Beijing 100071, China. yanbing72@hotmail.com
- Publication Type:Journal Article
- MeSH:
Animals;
Antibodies;
administration & dosage;
genetics;
metabolism;
Antibody Formation;
Base Sequence;
Blood-Brain Barrier;
drug effects;
Drug Delivery Systems;
Genetic Vectors;
Molecular Sequence Data;
Rats;
Rats, Sprague-Dawley;
Receptors, Transferrin;
immunology
- From:
Chinese Journal of Biotechnology
2004;20(3):342-347
- CountryChina
- Language:Chinese
-
Abstract:
The single-chain antibody gene (ox26-scFv) to transferrin receptor (TfR) was synthesized and amplified by three-step PCR. After sequencing, the gene was cloned into prokaryotic expression vector pTIG-Trx which carried thioredoxin (Trx) gene and a C-terminal His.tag. The Ox26-scFv proteins achieved 31% yields of total bacteria proteins at 20 degrees C, after 0.02mM IPTG induction using the strain E. coli BL21 (DE3). The soluble scFv proteins in cytoplasm suspension were about 35% and the inclusion bodies were about 65%. The soluble products were purified by immobilized metal chelation affinity chromatography (Ni-NTA), a single band with molecular weight 29 kD appeared on SDS-PAGE gel. Rat GH3 cell immunocytochemistry staining showed that Ox26-scFv protein could recognize and bind to transferrin receptor. Injected SD rats with Ox26-scFv proteins by tail veins, the antibodies were detected from brain tissues specially on the brain capillaries 4 h later which indicate that Ox26-scFv proteins have a good target function to brain capillaries and can permeate the blood-brain barrier mediated by the transferrin receptors.
