Characterization and Use of Monoclonal Antibodies Directed Against Human Erythropoietin That Recognize Different Antigenic Determinants
We have established four hybridoma cells that produce monoclonal antibodies (MoAbs) R2, R4, R6, and R12 directed toward recombinant human erythropoietin (rHuEPO). MoAbs R2, R4, and R6 bound to EPO with high affinities (kd = ∼2, 4, and 1 nmol/L, respectively) but MoAb R12 had a low affinity (240 nmol...
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| Veröffentlicht in: | Blood 1989-09, Vol.74 (4), p.1415-1423 |
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| creator | Goto, Masaaki Murakami, Akihiko Akai, Kunihisa Kawanishi, Gosei Ueda, Masatsugu Chiba, Hideo Sasaki, Ryuzo |
| description | We have established four hybridoma cells that produce monoclonal antibodies (MoAbs) R2, R4, R6, and R12 directed toward recombinant human erythropoietin (rHuEPO). MoAbs R2, R4, and R6 bound to EPO with high affinities (kd = ∼2, 4, and 1 nmol/L, respectively) but MoAb R12 had a low affinity (240 nmol/L). These antibodies inhibited the biological activity of rHuEPO and EPOs from humans, rats, mice, and rabbits. This inhibition was due to the blocking of EPO binding to the target cells. The fully deglycosylated rHuEPO bound to the MoAbs, indicating that they recognized peptide sequences of the antigen but not the carbohydrates attached to the antigen. An immunosorbent column with the immobilized MoAb R2 was effective for the rapid purification of EPO. MoAb R6 bound to EPO at a site(s) different from those to which other MoAbs bound. Based on this finding, a sensitive and rapid enzyme-linked immunosorbent assay of EPO, in which EPO was sandwiched between two MoAbs (R2 and R6), was developed. The assay measured plasma levels of EPO as low as 5 mU/mL within several hours. |
| doi_str_mv | 10.1182/blood.V74.4.1415.1415 |
| format | Article |
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MoAbs R2, R4, and R6 bound to EPO with high affinities (kd = ∼2, 4, and 1 nmol/L, respectively) but MoAb R12 had a low affinity (240 nmol/L). These antibodies inhibited the biological activity of rHuEPO and EPOs from humans, rats, mice, and rabbits. This inhibition was due to the blocking of EPO binding to the target cells. The fully deglycosylated rHuEPO bound to the MoAbs, indicating that they recognized peptide sequences of the antigen but not the carbohydrates attached to the antigen. An immunosorbent column with the immobilized MoAb R2 was effective for the rapid purification of EPO. MoAb R6 bound to EPO at a site(s) different from those to which other MoAbs bound. Based on this finding, a sensitive and rapid enzyme-linked immunosorbent assay of EPO, in which EPO was sandwiched between two MoAbs (R2 and R6), was developed. The assay measured plasma levels of EPO as low as 5 mU/mL within several hours.</description><identifier>ISSN: 0006-4971</identifier><identifier>EISSN: 1528-0020</identifier><identifier>DOI: 10.1182/blood.V74.4.1415.1415</identifier><identifier>PMID: 2475190</identifier><language>eng</language><publisher>Washington, DC: Elsevier Inc</publisher><subject>Analytical, structural and metabolic biochemistry ; Animals ; Antibodies, Monoclonal - immunology ; Antibodies, Monoclonal - physiology ; Antigen-Antibody Reactions ; Binding Sites, Antibody ; Binding, Competitive ; Biological and medical sciences ; Enzyme-Linked Immunosorbent Assay ; Epitopes - immunology ; Erythropoietin - immunology ; Erythropoietin - isolation & purification ; Erythropoietin - metabolism ; Female ; Fundamental and applied biological sciences. Psychology ; Glycoproteins ; Glycosylation ; Humans ; Mice ; Mice, Inbred BALB C ; Neutralization Tests ; Proteins ; Receptors, Cell Surface - analysis ; Receptors, Erythropoietin ; Recombinant Proteins - isolation & purification</subject><ispartof>Blood, 1989-09, Vol.74 (4), p.1415-1423</ispartof><rights>1989 American Society of Hematology</rights><rights>1990 INIST-CNRS</rights><lds50>peer_reviewed</lds50><oa>free_for_read</oa><woscitedreferencessubscribed>false</woscitedreferencessubscribed><citedby>FETCH-LOGICAL-c436t-a2881e3b402b6c752a8fefcda7a1ea8cd7f725eaba008c2d569d1ea9692889c43</citedby></display><links><openurl>$$Topenurl_article</openurl><openurlfulltext>$$Topenurlfull_article</openurlfulltext><thumbnail>$$Tsyndetics_thumb_exl</thumbnail><link.rule.ids>314,776,780,27901,27902</link.rule.ids><backlink>$$Uhttp://pascal-francis.inist.fr/vibad/index.php?action=getRecordDetail&idt=6674910$$DView record in Pascal Francis$$Hfree_for_read</backlink><backlink>$$Uhttps://www.ncbi.nlm.nih.gov/pubmed/2475190$$D View this record in MEDLINE/PubMed$$Hfree_for_read</backlink></links><search><creatorcontrib>Goto, Masaaki</creatorcontrib><creatorcontrib>Murakami, Akihiko</creatorcontrib><creatorcontrib>Akai, Kunihisa</creatorcontrib><creatorcontrib>Kawanishi, Gosei</creatorcontrib><creatorcontrib>Ueda, Masatsugu</creatorcontrib><creatorcontrib>Chiba, Hideo</creatorcontrib><creatorcontrib>Sasaki, Ryuzo</creatorcontrib><title>Characterization and Use of Monoclonal Antibodies Directed Against Human Erythropoietin That Recognize Different Antigenic Determinants</title><title>Blood</title><addtitle>Blood</addtitle><description>We have established four hybridoma cells that produce monoclonal antibodies (MoAbs) R2, R4, R6, and R12 directed toward recombinant human erythropoietin (rHuEPO). MoAbs R2, R4, and R6 bound to EPO with high affinities (kd = ∼2, 4, and 1 nmol/L, respectively) but MoAb R12 had a low affinity (240 nmol/L). These antibodies inhibited the biological activity of rHuEPO and EPOs from humans, rats, mice, and rabbits. This inhibition was due to the blocking of EPO binding to the target cells. The fully deglycosylated rHuEPO bound to the MoAbs, indicating that they recognized peptide sequences of the antigen but not the carbohydrates attached to the antigen. An immunosorbent column with the immobilized MoAb R2 was effective for the rapid purification of EPO. MoAb R6 bound to EPO at a site(s) different from those to which other MoAbs bound. Based on this finding, a sensitive and rapid enzyme-linked immunosorbent assay of EPO, in which EPO was sandwiched between two MoAbs (R2 and R6), was developed. The assay measured plasma levels of EPO as low as 5 mU/mL within several hours.</description><subject>Analytical, structural and metabolic biochemistry</subject><subject>Animals</subject><subject>Antibodies, Monoclonal - immunology</subject><subject>Antibodies, Monoclonal - physiology</subject><subject>Antigen-Antibody Reactions</subject><subject>Binding Sites, Antibody</subject><subject>Binding, Competitive</subject><subject>Biological and medical sciences</subject><subject>Enzyme-Linked Immunosorbent Assay</subject><subject>Epitopes - immunology</subject><subject>Erythropoietin - immunology</subject><subject>Erythropoietin - isolation & purification</subject><subject>Erythropoietin - metabolism</subject><subject>Female</subject><subject>Fundamental and applied biological sciences. Psychology</subject><subject>Glycoproteins</subject><subject>Glycosylation</subject><subject>Humans</subject><subject>Mice</subject><subject>Mice, Inbred BALB C</subject><subject>Neutralization Tests</subject><subject>Proteins</subject><subject>Receptors, Cell Surface - analysis</subject><subject>Receptors, Erythropoietin</subject><subject>Recombinant Proteins - isolation & purification</subject><issn>0006-4971</issn><issn>1528-0020</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>1989</creationdate><recordtype>article</recordtype><sourceid>EIF</sourceid><recordid>eNqFUU1vEzEQtRCohMJPqOQD4rbBdrwfPqEoLRSpCAm1XK1ZezYx2rWD7SC1f4C_jbOJeuUyc3gfM3qPkCvOlpx34mM_hmCXP1u5lEsueT2PF2TBa9FVjAn2kiwYY00lVctfkzcp_WKMy5WoL8iFkG3NFVuQv5sdRDAZo3uC7IKn4C19SEjDQL8FH8wYPIx07bPrg3WY6LWLWASWrrfgfMr09jCBpzfxMe9i2AeH2Xl6v4NMf6AJW--esIiGASP6PDtt0TtDr7GcnZwHn9Nb8mqAMeG7874kD59v7je31d33L18367vKyFWTKxBdx3HVSyb6xrS1gG7AwVhogSN0xrZDK2qEHhjrjLB1o2wBVKOKUBWPS_Lh5LuP4fcBU9aTSwbHETyGQ9Kt4o1a1V0h1ieiiSGliIPeRzdBfNSc6WMBei5AlwK01Mfs51F0V-cDh35C-6w6J17w92cckoFxiOCNS8-0pmml4kfapxMNSxh_HEadjENv0M7paxvcfx75BwLXqGI</recordid><startdate>19890901</startdate><enddate>19890901</enddate><creator>Goto, Masaaki</creator><creator>Murakami, Akihiko</creator><creator>Akai, Kunihisa</creator><creator>Kawanishi, Gosei</creator><creator>Ueda, Masatsugu</creator><creator>Chiba, Hideo</creator><creator>Sasaki, Ryuzo</creator><general>Elsevier Inc</general><general>The Americain Society of Hematology</general><scope>6I.</scope><scope>AAFTH</scope><scope>IQODW</scope><scope>CGR</scope><scope>CUY</scope><scope>CVF</scope><scope>ECM</scope><scope>EIF</scope><scope>NPM</scope><scope>AAYXX</scope><scope>CITATION</scope><scope>7X8</scope></search><sort><creationdate>19890901</creationdate><title>Characterization and Use of Monoclonal Antibodies Directed Against Human Erythropoietin That Recognize Different Antigenic Determinants</title><author>Goto, Masaaki ; Murakami, Akihiko ; Akai, Kunihisa ; Kawanishi, Gosei ; Ueda, Masatsugu ; Chiba, Hideo ; Sasaki, Ryuzo</author></sort><facets><frbrtype>5</frbrtype><frbrgroupid>cdi_FETCH-LOGICAL-c436t-a2881e3b402b6c752a8fefcda7a1ea8cd7f725eaba008c2d569d1ea9692889c43</frbrgroupid><rsrctype>articles</rsrctype><prefilter>articles</prefilter><language>eng</language><creationdate>1989</creationdate><topic>Analytical, structural and metabolic biochemistry</topic><topic>Animals</topic><topic>Antibodies, Monoclonal - immunology</topic><topic>Antibodies, Monoclonal - physiology</topic><topic>Antigen-Antibody Reactions</topic><topic>Binding Sites, Antibody</topic><topic>Binding, Competitive</topic><topic>Biological and medical sciences</topic><topic>Enzyme-Linked Immunosorbent Assay</topic><topic>Epitopes - immunology</topic><topic>Erythropoietin - immunology</topic><topic>Erythropoietin - isolation & purification</topic><topic>Erythropoietin - metabolism</topic><topic>Female</topic><topic>Fundamental and applied biological sciences. Psychology</topic><topic>Glycoproteins</topic><topic>Glycosylation</topic><topic>Humans</topic><topic>Mice</topic><topic>Mice, Inbred BALB C</topic><topic>Neutralization Tests</topic><topic>Proteins</topic><topic>Receptors, Cell Surface - analysis</topic><topic>Receptors, Erythropoietin</topic><topic>Recombinant Proteins - isolation & purification</topic><toplevel>peer_reviewed</toplevel><toplevel>online_resources</toplevel><creatorcontrib>Goto, Masaaki</creatorcontrib><creatorcontrib>Murakami, Akihiko</creatorcontrib><creatorcontrib>Akai, Kunihisa</creatorcontrib><creatorcontrib>Kawanishi, Gosei</creatorcontrib><creatorcontrib>Ueda, Masatsugu</creatorcontrib><creatorcontrib>Chiba, Hideo</creatorcontrib><creatorcontrib>Sasaki, Ryuzo</creatorcontrib><collection>ScienceDirect Open Access Titles</collection><collection>Elsevier:ScienceDirect:Open Access</collection><collection>Pascal-Francis</collection><collection>Medline</collection><collection>MEDLINE</collection><collection>MEDLINE (Ovid)</collection><collection>MEDLINE</collection><collection>MEDLINE</collection><collection>PubMed</collection><collection>CrossRef</collection><collection>MEDLINE - Academic</collection><jtitle>Blood</jtitle></facets><delivery><delcategory>Remote Search Resource</delcategory><fulltext>fulltext</fulltext></delivery><addata><au>Goto, Masaaki</au><au>Murakami, Akihiko</au><au>Akai, Kunihisa</au><au>Kawanishi, Gosei</au><au>Ueda, Masatsugu</au><au>Chiba, Hideo</au><au>Sasaki, Ryuzo</au><format>journal</format><genre>article</genre><ristype>JOUR</ristype><atitle>Characterization and Use of Monoclonal Antibodies Directed Against Human Erythropoietin That Recognize Different Antigenic Determinants</atitle><jtitle>Blood</jtitle><addtitle>Blood</addtitle><date>1989-09-01</date><risdate>1989</risdate><volume>74</volume><issue>4</issue><spage>1415</spage><epage>1423</epage><pages>1415-1423</pages><issn>0006-4971</issn><eissn>1528-0020</eissn><abstract>We have established four hybridoma cells that produce monoclonal antibodies (MoAbs) R2, R4, R6, and R12 directed toward recombinant human erythropoietin (rHuEPO). MoAbs R2, R4, and R6 bound to EPO with high affinities (kd = ∼2, 4, and 1 nmol/L, respectively) but MoAb R12 had a low affinity (240 nmol/L). These antibodies inhibited the biological activity of rHuEPO and EPOs from humans, rats, mice, and rabbits. This inhibition was due to the blocking of EPO binding to the target cells. The fully deglycosylated rHuEPO bound to the MoAbs, indicating that they recognized peptide sequences of the antigen but not the carbohydrates attached to the antigen. An immunosorbent column with the immobilized MoAb R2 was effective for the rapid purification of EPO. MoAb R6 bound to EPO at a site(s) different from those to which other MoAbs bound. Based on this finding, a sensitive and rapid enzyme-linked immunosorbent assay of EPO, in which EPO was sandwiched between two MoAbs (R2 and R6), was developed. The assay measured plasma levels of EPO as low as 5 mU/mL within several hours.</abstract><cop>Washington, DC</cop><pub>Elsevier Inc</pub><pmid>2475190</pmid><doi>10.1182/blood.V74.4.1415.1415</doi><tpages>9</tpages><oa>free_for_read</oa></addata></record> |
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| source | MEDLINE; Elektronische Zeitschriftenbibliothek - Frei zugängliche E-Journals; Alma/SFX Local Collection |
| subjects | Analytical, structural and metabolic biochemistry Animals Antibodies, Monoclonal - immunology Antibodies, Monoclonal - physiology Antigen-Antibody Reactions Binding Sites, Antibody Binding, Competitive Biological and medical sciences Enzyme-Linked Immunosorbent Assay Epitopes - immunology Erythropoietin - immunology Erythropoietin - isolation & purification Erythropoietin - metabolism Female Fundamental and applied biological sciences. Psychology Glycoproteins Glycosylation Humans Mice Mice, Inbred BALB C Neutralization Tests Proteins Receptors, Cell Surface - analysis Receptors, Erythropoietin Recombinant Proteins - isolation & purification |
| title | Characterization and Use of Monoclonal Antibodies Directed Against Human Erythropoietin That Recognize Different Antigenic Determinants |
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