Immobilisation of monocytes to a solid support: a model for the study of ligand-binding interactions and plasminogen activation at the cell surface.
The functional and immunological identification of receptors expressed by cells of the monocyte/ macrophage lineage may be facilitated with the use of immobilised cells. A procedure is described here for attaching human blood monocytes, alveolar macrophages, and THP-1 cells to a solid support activated with polymerised glutaraldehyde. Homogeneous monolayers visualised by optical microscopy were obtained at predefined input cell densities and were quantitatively characterised with the use of 125I-plasminogen (35000+/-2772 cells/well at approximately 76000 cells/50 microL) and 125I-pro-urokinase (39000+/-3839 cells/well at approximately 86000 cells/50 microL). The cells remained stably attached during washing and incubation procedures in ligand-binding studies. The functionality of membrane receptors and acceptors of the immobilised cells for a number of ligands was verified. Parameters of the interaction of plasminogen, urokinase, and human immunoglobulin G with their corresponding receptors were similar to those previously reported using cells in suspension. The functionality of bound ligands, such as urokinase and plasminogen, was verified by measuring their ability to generate plasmin. We conclude that immobilised monocytes/macrophages are a useful tool for studying ligand interactions with membrane proteins and for the realisation of plasminogen activation studies at the surface of the cell membrane.
Soulat
T
T
INSERM U.143 Hémostase-Biologie Vasculaire, Hôpital de Bicêtre, Paris, France.
Dominguez
M
M
Martin
I
I
Anglés-Cano
E
E
D002452:Cell Count; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism*; D018914:Cells, Immobilized; D005976:Glutaral; D017261:Glycosylphosphatidylinositols / Q000378:metabolism; D006801:Humans; D008024:Ligands; D016676:Macrophages, Alveolar / Q000166:cytology / Q000378:metabolism; D009000:Monocytes / Q000166:cytology / Q000378:metabolism*; D010958:Plasminogen / Q000378:metabolism*; D011485:Protein Binding; D011956:Receptors, Cell Surface / Q000378:metabolism*; D017452:Receptors, IgG / Q000378:metabolism; D014568:Urokinase-Type Plasminogen Activator / Q000378:metabolism
D017261:Glycosylphosphatidylinositols; D008024:Ligands; D011956:Receptors, Cell Surface; D017452:Receptors, IgG; D010958:Plasminogen; D014568:Urokinase-Type Plasminogen Activator; D005976:Glutaral