Preparation and biological properties of monoclonal antibodies against human erythropoietin.
Several monoclonal antibodies (Mabs) against human erythropoietin (H-EPO) were obtained by fusion of myeloma cells with splenocytes from mice immunized with purified recombinant H-EPO. Three Mabs (D7, E14 and E73) were selected by radioimmunoprecipitation on the basis of their high affinity against H-EPO. Their dissociation constants were 0.3 nM for E14 and D7, and 17 nM for E73. The immunochemical properties of these Mabs were analyzed in respect to their capacity to react, to purify, and to inhibit the biological activities of H-EPO. a) detection by Western blotting techniques: among the 3 Mabs only D7 was effective by this technique. b) purification: the best results were observed with E14, an approximately 200-fold purification of H-EPO from culture supernatants was obtained in a single immunopurification step. c) inhibition of the biological activity: the specific binding of 125I-labelled H-EPO to its cellular receptor was inhibited by E14 and E73 but not by D7. These three Mabs exhibited similar effects as far as the inhibition of the proliferation of H-EPO dependent cells (measured by 3H-thymidine incorporation) was concerned.
Ben Ghanem
A
A
Institut National de Transfusion Sanguine, Paris, France.
Pallu
S
S
Winchenne
J J
JJ
Raymonjean
N
N
Mayeux
P
P
Goossens
D
D
Boffa
G A
GA
Casadevall
N
N
Cartron
J P
JP
Lambin
P
P
D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology / Q000302:isolation & purification*; D000915:Antibody Affinity; D000918:Antibody Specificity; D004921:Erythropoietin / Q000037:antagonists & inhibitors / Q000276:immunology* / Q000378:metabolism; D006801:Humans; D006825:Hybridomas / Q000276:immunology; D007132:Immunoglobulin Isotypes / Q000276:immunology / Q000302:isolation & purification; D066298:In Vitro Techniques; D051379:Mice; D008807:Mice, Inbred BALB C; D009500:Neutralization Tests; D011817:Rabbits; D017467:Receptors, Erythropoietin / Q000378:metabolism
D000911:Antibodies, Monoclonal; D007132:Immunoglobulin Isotypes; D017467:Receptors, Erythropoietin; D004921:Erythropoietin