Identification of high affinity receptors for human monocyte chemoattractant protein-1 on human monocytes.
The binding of human monocyte chemoattractant protein-1 (MCP-1) to human monocytes was studied. MCP-1 was radioiodinated with Iodo-beads (Pierce Chemical Co., Rockford, IL) without significant loss of biologic activity. 125I-MCP-1 binding to PBMC occurred within 5 min at 0 degrees C and the binding was inhibited by unlabeled MCP-1 dose dependently but not by neutrophil attractant/activation protein-1 or FMLP. 125I-MCP-1 bound to monocytes; no significant binding to either neutrophils or lymphocytes was observed. Scatchard plot analysis indicated that monocytes had a minimum of 1700 +/- 600 binding sites per cell with a Kd of 1.9 +/- 0.2 x 10(-9) M. For analysis of binding by flow cytometry, MCP-1 was biotinylated. In contrast to radioiodination, biotinylation resulted in loss of activity; potency was 10-fold less, but the efficacy was retained. Detection by flow cytometry of bound biotinylated MCP-1 with avidin-FITC confirmed results obtained with 125I-MCP-1. Biotinylated MCP-1 bound to monocytes but not to lymphocytes; and the binding was inhibited by a 100-fold excess of unlabeled MCP-1.
Yoshimura
T
T
Immunopathology Section, National Cancer Institute-Frederick Cancer Research Facility, MD 21701.
Leonard
E J
EJ
D018932:Chemokine CCL2; D002630:Chemotactic Factors / Q000502:physiology*; D002634:Chemotaxis, Leukocyte; D005434:Flow Cytometry; D006801:Humans; D066298:In Vitro Techniques; D008214:Lymphocytes / Q000378:metabolism; D008970:Molecular Weight; D009000:Monocytes / Q000502:physiology*; D011956:Receptors, Cell Surface / Q000378:metabolism*
D018932:Chemokine CCL2; D002630:Chemotactic Factors; D011956:Receptors, Cell Surface