Unidirectional potentiation of binding between two anti-FBP MAbs: evaluation of the involved mechanisms.
The monoclonal antibody MOv19 directed to a folate binding protein shows temperature-dependent potentiation of binding of the noncompeting monoclonal antibody MOv18 to the relevant antigen, but the mechanism involved in this phenomenon had remained unclear. Use of chimeric versions of both monoclonal antibodies and the F(ab')2 and Fab fragments of MOv19 revealed an increment in MOv18 binding in all combinations irrespective of the origin of the Fc portion of the monoclonal antibody. The potentiating effect of bivalent MOv19 fragments on 125I-MOv18 binding was similar to that of the entire monoclonal antibody and occurred at saturating concentrations of both reagents at which monovalent binding prevails. Similarly, the monovalent fragment also induced a significant increase in MOv18 binding. However, the potentiation occurred only at very high concentrations of antibody fragment. Homologous inhibition was drastically reduced using MOv19 Fab fragment, suggesting a low binding stability of the monovalent reagent. Immunoblotting analysis and binding in the presence of exogenous purified folate binding protein indicated a cross-linking between soluble and cell surface molecules mediated by the bivalent monoclonal antibodies. The extent of the increase in MOv18 binding at 0 degrees C with high amounts of exogenous folate binding protein was lower than that obtained at 37 degrees C in the absence of added molecule. Release of 125I-MOv18 from the cell surface was significantly higher in the absence of MOv19 than in its presence.(ABSTRACT TRUNCATED AT 250 WORDS)
Casalini
P
P
Division of Experimental Oncology E, Istituto Nazionale Tumori, Milano, Italy.
Mezzanzanica
D
D
Valota
O
O
Adobati
E
E
Tomassetti
A
A
Colnaghi
M I
MI
Canevari
S
S
D000911:Antibodies, Monoclonal / Q000737:chemistry / Q000276:immunology*; D000915:Antibody Affinity; D000937:Antigen-Antibody Reactions; D001666:Binding Sites, Antibody; D001667:Binding, Competitive; D002277:Carcinoma; D002352:Carrier Proteins / Q000276:immunology*; D005260:Female; D058973:Folate Receptors, GPI-Anchored; D006801:Humans; D015151:Immunoblotting; D007140:Immunoglobulin Fab Fragments / Q000276:immunology; D007700:Kinetics; D010051:Ovarian Neoplasms; D011487:Protein Conformation; D011956:Receptors, Cell Surface / Q000276:immunology*; D011993:Recombinant Fusion Proteins / Q000276:immunology; D013997:Time Factors; D014407:Tumor Cells, Cultured
D000911:Antibodies, Monoclonal; D002352:Carrier Proteins; D058973:Folate Receptors, GPI-Anchored; D007140:Immunoglobulin Fab Fragments; D011956:Receptors, Cell Surface; D011993:Recombinant Fusion Proteins