Platelet-derived growth factor labeled to colloidal gold for use as a mitogenic receptor probe.
Studies by others utilizing 125I-PDGF have indicated that target cells express a high affinity surface receptor for PDGF. We have bound purified platelet-derived growth factor (PDGF) to gold colloid particles to explore the interaction of PDGF with mouse 3T3 cells. The gold-PDGF complex consists of approximately 26 PDGF molecules electrostatically absorbed to gold colloid (approximately 14.1 nm). The gold-PDGF complex induced mitogenic stimulation similar to unbound PDGF, although a 5 to 6 fold greater amount of complexed PDGF was required for the same effect. Incubation of the gold-PDGF complex with 3T3 cells for 4 h at 4 degrees C revealed that 98% of the membrane binding was randomly distributed on the cell surface with respect to coated pits, with each cell binding 7000 to 11000 complexes. Addition of a 20-fold excess of unlabeled PDGF reduced surface binding of the gold-PDGF complex by 87% (1230 probes/cell). Warming to 37 degrees C followed by time-interval fixation permitted visualization of endocytosis of the complexes in coated vesicles (1-3 min), internalization (3-15 min) and lysosomal accumulation (15-60 min). Pretreatment of cultures with monensin (2 h, 10 microM) abolished receptor binding, internalization and subsequent mitogenesis of the gold-PDGF complex. These studies support the suggestion that PDGF requires a surface receptor to elicit mitogenesis.
D000818:Animals; D002462:Cell Membrane / Q000648:ultrastructure; D002478:Cells, Cultured; D003102:Colloids; D006046:Gold; D051379:Mice; D008807:Mice, Inbred BALB C; D008854:Microscopy, Electron; D010982:Platelet-Derived Growth Factor; D011956:Receptors, Cell Surface / Q000032:analysis*; D011975:Receptors, Mitogen / Q000032:analysis*; D017479:Receptors, Platelet-Derived Growth Factor
D003102:Colloids; D010982:Platelet-Derived Growth Factor; D011956:Receptors, Cell Surface; D011975:Receptors, Mitogen; D006046:Gold; D017479:Receptors, Platelet-Derived Growth Factor