A preliminary optical and electron microscopic study of the beta(1) integrin distribution pattern of human osteosarcoma-derived cells.
Immunogold labelling was used to study the organisation of the beta(1) integrins on osteosarcoma-derived osteoblasts (Saos-2 and MG-63). Monolayers of cells were prepared in multiwell culture plates on both uncovered and collagen-covered coverslips, and beta(1) integrins were primarily labelled using mouse monoclonal antibodies to beta(1) integrins. Indirect immunofluorescence labels using an anti-mouse fluorescein-conjugated goat antibody showed an even distribution of the beta(1) integrins on the cell membranes of all cell types used. A concentration of 2 microg/ml of the primary antibodies and a 1:100 dilution of the secondary antibodies were determined as the optimal concentration for labelling to use with indirect localisation of the primary antibodies gold conjugated to goat anti-mouse antibodies and viewed under an electron microscope. Ten nanometre gold particles were used for transmission electron microscopy (TEM) and 40 nm gold particles for scanning electron microscopy. TEM showed that beta(1) integrins were mainly clustered on the cell membrane processes with less labelling on the cell membranes themselves. The distribution of beta(1) integrins on osteosarcoma cells supports the concept that integrins may function by forming focal adhesions at the site of the cytoplasmic membrane processes.
Banai
Kiarash
K
Department of Orthodontics, Floor 22, Guy's Tower, Guy's, King's and St. Thomas' Dental Institute, King's College London, SE1 9RT, London, UK.
Brady
Ken
K
McDonald
Fraser
F
D001859:Bone Neoplasms / Q000378:metabolism* / Q000473:pathology; D019084:Fluorescent Antibody Technique, Indirect; D006801:Humans; D019012:Integrin beta1 / Q000378:metabolism*; D008854:Microscopy, Electron; D010006:Osteoblasts / Q000378:metabolism* / Q000648:ultrastructure; D012516:Osteosarcoma / Q000378:metabolism* / Q000473:pathology; D013194:Staining and Labeling / Q000379:methods; D014407:Tumor Cells, Cultured
D019012:Integrin beta1