Immunolocalization of the lactotransferrin receptor on the human T lymphoblastic cell line Jurkat.
Monoclonal antibodies have been raised against the soluble lactotransferrin binding protein purified from the cell culture supernatant of Jurkat cell line, a human T-lymphoblastic cell. All monoclonal antibodies were able to specifically bind to the membrane of Jurkat cells. One of the monoclonal antibodies, DP5B3G10, recognized both the soluble lactotransferrin-binding protein and the membrane lymphocyte lactotransferrin receptor after SDS-PAGE in presence of 2-mercaptoethanol and electrotransfer on nitrocellulose. The monoclonal antibody DP5B3G10 inhibited the binding of lactotransferrin to Jurkat cells and human peripheral activated lymphocytes. In addition, lactotransferrin inhibited the binding of the monoclonal antibody to the cell surface. These results suggest that the 95 kDa lactotransferrin-binding protein isolated from the cell culture medium corresponds to the soluble form of the 105 kDa lymphocyte lactotransferrin receptor. Corresponding proteins of 105 kDa molecular mass were identified in Jurkat and CEM T-cells and Raji B-cells. Finally, the monoclonal antibody DP5B3G10 was used to immunolocalize the lactotransferrin receptor on the Jurkat cells. Using fluorescence and electron microscopy, the receptor was localized both inside and at the cell surface. The cell membrane receptor was associated into clusters. After permeabilization of the plasma membrane, the staining was positive in the peri-membrane area. The region near the nucleus was devoid of receptor.
Bi
B Y
BY
Laboratoire de Chimie Biologique, UMR no. 111 du CNRS, Université des Sciences et Technologies de Lille, Villeneuve d'Ascq/France.
Leveugle
B
B
Liu
J L
JL
Collard
A
A
Coppe
P
P
Roche
A C
AC
Nillesse
N
N
Capron
M
M
Spik
G
G
Mazurier
J
J
D000911:Antibodies, Monoclonal; D001667:Binding, Competitive; D002460:Cell Line; D003470:Culture Media; D005455:Fluorescent Antibody Technique; D006801:Humans; D007136:Immunoglobulins / Q000096:biosynthesis; D008854:Microscopy, Electron; D011956:Receptors, Cell Surface / Q000032:analysis*; D012995:Solubility; D013601:T-Lymphocytes / Q000737:chemistry*
D000911:Antibodies, Monoclonal; D003470:Culture Media; D007136:Immunoglobulins; D011956:Receptors, Cell Surface; C038340:lactoferrin receptors