Isolation and identification of a human single chain Fv antibody against amyloid-beta 1-42 soluble oligomers from a human phage display library / 生物工程学报
Chinese Journal of Biotechnology
;
(12): 1195-1203, 2009.
Artículo
en Chino
| WPRIM
| ID: wpr-296938
ABSTRACT
To get specific scFv (Single-chain fragment variable) antibody against soluble Abeta1-42(Amyloid-beta) oligomers, we constructed a human single-chain Fv (scFv) antibody library by phage display technology. Using RT-PCR, we amplified the variable heavy (VH) and variable light (VL) genes from peripheral blood lymphocytes (PBL). Then we obtained the scFv fragments through SOE-PCR, and the scFv fragments were cloned into the vector pCANTAB5E and electroporated into competent Escherichia coli TG1 cells. Consequently, a scFv phage display library containing 2.5 x 10(9) clones was constructed. The recombinant phagemids were rescued by reinfection of helper phage M13K07. Recombinant phages specific for Abeta1-42 oligomers were enriched after four rounds of biopanning and the antigen-positive clones were selected from the enriched clones by phage ELISA. Positive clone B19 was used to infect E. coli HB2151 to express soluble scFv antibody. SDS-PAGE and Western blotting analysis showed that the soluble scFv B19 antibody was expressed successfully and could bind specifically to Abeta1-42 trimer and protofiber. The specific scFv against Abeta1-42 oligomers can be used in the therapeutic research on Alzheimer's disease.
Texto completo:
Disponible
Índice:
WPRIM (Pacífico Occidental)
Asunto principal:
Fragmentos de Péptidos
/
Péptidos beta-Amiloides
/
Biblioteca de Péptidos
/
Alergia e Inmunología
/
Anticuerpos de Cadena Única
/
Genética
/
Especificidad de Anticuerpos
Tipo de estudio:
Estudio pronóstico
Límite:
Humanos
Idioma:
Chino
Revista:
Chinese Journal of Biotechnology
Año:
2009
Tipo del documento:
Artículo
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