Preparation of Fluorescent Recombinant Shiga Toxin B Subunit and Its Application to Flow Cytometry.
Methods Mol Biol
; 2132: 463-474, 2020.
Article
en En
| MEDLINE
| ID: mdl-32306353
Shiga toxin (Stx) is a major virulence factor of enterohemorrhagic Escherichia coli (E. coli). Stx consists of one enzymatic A subunit and five B subunits (StxB) that are involved in binding. The StxB pentamer specifically recognizes a glycosphingolipid, globotriaosylceramide (Gb3), as a receptor; therefore, it can be used as a probe to detect Gb3. This chapter describes the preparation of recombinant Stx1B proteins using E. coli, their conjugation with fluorescent dyes, and their application for flow cytometry. The prepared fluorescent StxB proteins bound to cells of several lines, including the HeLa human cervix adenocarcinoma cell line and the THP-1 human monocytic leukemia cell line. Furthermore, the probe was useful for confirmation of several sphingolipid-deficient HeLa cell lines that were constructed using genome editing.
Palabras clave
Texto completo:
1
Colección:
01-internacional
Base de datos:
MEDLINE
Asunto principal:
Rodaminas
/
Ácidos Sulfónicos
/
Toxinas Shiga
/
Escherichia coli Enterohemorrágica
Límite:
Humans
Idioma:
En
Revista:
Methods Mol Biol
Asunto de la revista:
BIOLOGIA MOLECULAR
Año:
2020
Tipo del documento:
Article
País de afiliación:
Japón
Pais de publicación:
Estados Unidos