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Immuno‐Dot Blot Analysis of a Monoclonal Antibody that Blocks the Human Monocyte Response to Migration Inhibitory Factor a
Author(s) -
LIU DAVID Y.,
III ROBERT F. TODD
Publication year - 1987
Publication title -
annals of the new york academy of sciences
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.712
H-Index - 248
eISSN - 1749-6632
pISSN - 0077-8923
DOI - 10.1111/j.1749-6632.1987.tb29531.x
Subject(s) - medical school , medicine , medical laboratory , library science , pathology , medical education , computer science
Mononuclear phagocytes are critical for both the specific and nonspecific functions of the immune system. One aspect is the communication that occurs between mononuclear phagocytes and lymphocytes. This interaction proceeds by way of soluble mediators secreted by these cells, such as migration inhibitory factor (MIF). MIF is produced and secreted by activated lymphocytes and inhibits the migration of mononuclear phagocytes. We have previously established that membrane glycolipids of mononuclear phagocytes play a significant role in the regulation of the human monocyte response to MIF.' Recently, a panel of murine monoclonal antibodies that recognize surface antigens specific for human cells of the mononuclear phagocyte lineage was developed by Todd and c0workers.2,~ These antibodies were used to further define the membrane components essential for the monocyte response to human MIF. We found that at least one monoclonal antibody (M03e) completely blocks the human monocyte response to MIF? This represents the first evidence for a monocyte-specific antigen that participates in a cellular function. The purpose of this study was to investigate by an immuno-dot analysis the nature of the antigen recognized by Mo3e. Cells isolated from peripheral blood or carried as transformed cell lines were lysed in Tris-saline buffer containing NP-40 detergent, protease inhibitors, and iodoacetamide. The lysates were then centrifuged, and the supernatants serially diluted before being applied to nitrocellulose paper as spots in a 96-well suction apparatus (Schleicher and Schuell, Keene, NH). After blocking nonspecific binding sites on the paper, it was incubated sequentially with Mo3e, rabbit anti-