A novel monoclonal antibody for detection of galectin-9 in tissue sections: application to human tissues infected by oncogenic viruses.
BACKGROUND
Galectin-9 is a mammalian lectin which possesses immunosuppressive properties. Excessive production of galectin-9 has been reported in two types of human virus-associated diseases chronic hepatitis C and nasopharyngeal carcinoma associated to the Epstein-Barr virus. The objective of this study was to produce new monoclonal antibodies targeting galectin-9 in order to improve its detection in clinical samples, especially on tissue sections analysed by immunohistochemistry.


METHODS
Hybridomas were produced through immunization of mice with the recombinant c-terminus part of galectin-9 (residues 191 to 355 of the long isoform) and semi-solid fusion of spleen cells with Sp2/0 cells. Monoclonal antibodies were characterized using ELISA, epitope mapping, western blot and immunohistochemistry.


RESULTS
We selected seven hybridomas producing antibodies reacting with our recombinant c-terminus galectin-9 in ELISA. Five of them reacted with the epitope "TPAIPPMMYPHPA" (common to all isoforms, residues 210 to 222 of the long isoform) and stained all three isoforms of galectin-9 analysed by western blot. One of them, 1G3,demonstrated very good sensitivity and specificity when used for immunohistochemistry. Using 1G3, we could confirm the intense and constant expression of galectin-9 by Epstein-Barr virus positive malignant cells from nasopharyngeal carcinomas. In most samples, specific staining was detected in both cytoplasm and nuclei. Galectin-9 was also detected in liver biopsies from patients infected by the human hepatitis C or B viruses with expression not only in inflammatory leucocytes and Kupffer cells, but also in hepatocytes. In contrast, galectin-9 was virtually absent in non-infected liver specimens.


CONCLUSIONS
The 1G3 monoclonal antibody will be a powerful tool to assess galectin-9 expression and distribution especially in diseases related to oncogenic viruses.
University Paris-Sud 11, CNRS-UMR 8126, Institut de Cancérologie Gustave Roussy, 114 rue Edouard Vaillant, 94805, Villejuif cedex, France. pierre.busson@igr.fr.
Clément
C
Barjon
Toshiro
T
Niki
Benjamin
B
Vérillaud
Paule
P
Opolon
Pierre
P
Bedossa
Mitsuomi
M
Hirashima
Stéphanie
S
Blanchin
Michel
M
Wassef
Hugo R
HR
Rosen
Anne-Sophie
AS
Jimenez
Ming
M
Wei
Pierre
P
Busson