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Production and Evaluation of Immunoreactivity of Poly Lysine-Tagged Single Chain Fragment Variable (ScFv) Lym-1 Antibody for Direct Conjugation to Fluorescence Dye / 핵의학분자영상
Nuclear Medicine and Molecular Imaging ; : 487-494, 2009.
Article Dans Coréen | WPRIM | ID: wpr-155609
ABSTRACT

PURPOSE:

Small size of recombinant scFv antibody has many advantages such as rapid blood clearances and improved targeting antibodies to tumor region. On the other hand owing to small size, number of amino group is insufficient in conjugation with chelator and fluorescence labeling. This study is to introduce poly lysine tag to the C-terminal end of scFv lym-1 sequence for fluorescence chelator conjugation. MATERIALS AND

METHODS:

Poly lysine scFv lym-1 gene, cloned into pET-22b (+) vector, was expressed in E. coli BL21 (DE3) strain. Antibody purification was performed with Ni-NTA column and then size exclusion column chromatography. Expression and purification levels of poly lysine tagged scFv lym-1 antibody were confirmed by western blot analysis. I-124, I-125, I-131 and Tc-99m were used for radiolabeling of purified poly lysine scFv lym-1. Flow cytometry analysis of FITC conjugated poly lysine scFv lym-1 was performed for confirmation of immunoreactivity of human Burkitt`s lymphoma cells.

RESULTS:

Poly lysine scFv lym-1 antibody was purified through two steps and identified as molecular weight of 48 KDa. Radiolabeling yields of I-124, I-125, I-131 and Tc-99m into poly lysine scFv lym-1 were >99%, >99%, >95% and >99%, respectively. Flow cytometry analysis of poly lysine scFv and scFv lym-1 was showed similar immunoreactivity to human Burkitt`s lymphoma cells.

CONCLUSION:

Poly lysine tag was useful for the sufficient number of amino groups to scFv lym-1 antibody for chelator conjugation with minimizing loss of immunoreactivity.
Sujets)

Texte intégral: Disponible Indice: WPRIM (Pacifique occidental) Sujet Principal: Entorses et foulures / Technique de Western / Chromatographie / Fluorescéine-5-isothiocyanate / Clones cellulaires / Cytométrie en flux / Fluorescence / Main / Lymphomes / Lysine Type d'étude: Étude pronostique Limites du sujet: Humains langue: Coréen Texte intégral: Nuclear Medicine and Molecular Imaging Année: 2009 Type: Article

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Texte intégral: Disponible Indice: WPRIM (Pacifique occidental) Sujet Principal: Entorses et foulures / Technique de Western / Chromatographie / Fluorescéine-5-isothiocyanate / Clones cellulaires / Cytométrie en flux / Fluorescence / Main / Lymphomes / Lysine Type d'étude: Étude pronostique Limites du sujet: Humains langue: Coréen Texte intégral: Nuclear Medicine and Molecular Imaging Année: 2009 Type: Article