The Experts below are selected from a list of 2532 Experts worldwide ranked by ideXlab platform
Jeanmichel Saveant - One of the best experts on this subject based on the ideXlab platform.
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activation and diffusion in the kinetics of adsorption and molecular recognition on surfaces enzyme amplified electrochemical approach to biorecognition dynamics illustrated by the binding of antibodies to Immobilized antigens
Journal of the American Chemical Society, 1999Co-Authors: Christian Bourdillon, Christophe Demaille, Jacques Moiroux, Jeanmichel SaveantAbstract:An electrochemical method is proposed for investigating the dynamics of recognition between a biomolecule and an Immobilized receptor. It involves redox labeling of the solute molecule and monitoring the binding by the electrochemical response of the electrode onto which the receptor is Immobilized. With large biomolecules, as, for example, antigens and antibodies, leading to small surface concentrations, simple redox labeling may prove insufficient to obtain detectable responses. Redox enzymes are then advantageously used as labels thanks to the signal amplification offered by their catalytic properties. The applicability of the method is illustrated by the reaction of an Immobilized monolayer of goat IgG antigen (or of one Fab fragment) with an antigoat antibody labeled with glucose oxidase. Particular care is taken to free the kinetic data from the effect of diffusion. The latter factor may interfere whatever the detection technique. A full account of the combination between recognition kinetics and di...
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activation and diffusion in the kinetics of adsorption and molecular recognition on surfaces enzyme amplified electrochemical approach to biorecognition dynamics illustrated by the binding of antibodies to Immobilized antigens
Journal of the American Chemical Society, 1999Co-Authors: Christian Bourdillon, Christophe Demaille, Jacques Moiroux, Jeanmichel SaveantAbstract:An electrochemical method is proposed for investigating the dynamics of recognition between a biomolecule and an Immobilized receptor. It involves redox labeling of the solute molecule and monitori...
D A Dmitriev - One of the best experts on this subject based on the ideXlab platform.
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kinetic analysis of interactions between bispecific monoclonal antibodies and Immobilized antigens using a resonant mirror biosensor
Journal of Immunological Methods, 2003Co-Authors: D A Dmitriev, Yulia S Massino, O L SegalAbstract:A resonant mirror biosensor (IAsys) protocol is described for the comparative kinetic analysis of the ability of monoclonal antibodies (Mabs) and bispecific antibodies (Babs) to bind Immobilized antigens. The protocol has been optimized and validated using the panel of affinity-purified antibodies, including two parental Mabs, one specific to human immunoglobulin G (hIgG) and another specific to horseradish peroxidase (HRP), and a Bab derived thereof by cell fusion (anti-hIgG/HRP Bab). The real-time kinetic analysis of antigen-antibody interactions using this protocol allows to demonstrate the differences in the avidity of bivalently binding Mabs and monovalent Babs. As shown in our previous study [J. Immunol. Methods 261 (2002) 103], the observed equilibrium association constants (Kass) determined by IAsys using this protocol yield figures almost overlapping with those obtained by solid-phase radioimmunoassay (RIA). The described protocol is suited for the investigation of the effects of valency on the binding properties of antibodies. It also may be applied for the selection of Mabs and Babs with desired features, for different fields of application.
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analysis of bispecific monoclonal antibody binding to Immobilized antigens using an optical biosensor
Biochemistry, 2002Co-Authors: D A Dmitriev, Yulia S Massino, O L Segal, M Smirnova, E V Pavlova, G I Kolyaskina, K G Gurevich, O V Gnedenko, Yuri D Ivanov, A I ArchakovAbstract:The interaction between two different monoclonal antibodies (Mabs) and their corresponding bispecific antibodies (Babs) with Immobilized antigens was investigated using an optical biosensor (IAsys). The analyzed panel of affinity-purified antibodies included two parental Mabs (one of which was specific to human IgG (hIgG), and another one to horseradish peroxidase (HRP)), as well as Babs derived thereof (anti-hIgG/HRP). Babs resulting from the fusion of parental hybridomas bear two antigen-binding sites toward two different antigens and thus may interact with Immobilized antigen through only one antigen-binding site (monovalently). Using an IAsys biosensor this study shows that the bivalent binding of Mabs predominates over the monovalent binding with Immobilized HRP, whereas anti-hIgG parental Mabs were bound monovalently to the Immobilized hIgG. The observed equilibrium association constant (K(ass)) values obtained in our last work [1] by solid-phase radioimmunoassay are consistent with those constants obtained by IAsys. The K(ass) of anti-HRP Mabs was about 50 times higher than that of anti-HRP shoulder of Babs. The dissociation rate constant (k(diss)) for anti-HRP shoulder of Babs was 21 times higher than k(diss) for anti-HRP Mabs. The comparison of the kinetic parameters for bivalent anti-HRP Mabs and Babs derived from anti-Mb/HRP and anti-hIgG/HRP, allowed to calculate that 95% of bound anti-HRP Mabs are bivalently linked with Immobilized HRP, whereas only 5% of bound anti-HRP Mabs are monovalently linked. In general, the data obtained indicate that Babs bearing an enzyme-binding site may not be efficiently used instead of traditional antibody-enzyme conjugates in the case of binding of bivalent Mabs.
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analysis of the binding of bispecific monoclonal antibodies with Immobilized antigens human igg and horseradish peroxidase using a resonant mirror biosensor
Journal of Immunological Methods, 2002Co-Authors: D A Dmitriev, Yulia S Massino, O L Segal, M Smirnova, E V Pavlova, G I Kolyaskina, K G Gurevich, O V Gnedenko, Yuri D Ivanov, A I ArchakovAbstract:Abstract The interaction between two monoclonal antibodies (mAbs) and their corresponding bispecific antibody (bAb) with Immobilized antigens has been examined using a resonant mirror biosensor (IAsys). BAbs were produced by cell fusion. The analysed panel of affinity-purified antibodies included two parental mAbs, one specific to human IgG (hIgG), and another specific to horseradish peroxidase (HRP), and a bAb derived thereof (anti-hIgG/HRP). The real-time analysis showed the drastic differences in the avidity of bivalent anti-HRP mAbs and anti-HRP shoulder of bAbs. Thus, the observed equilibrium association constant ( K ass ) of anti-HRP mAbs was about 50 times higher that of anti-HRP shoulder of bAbs. The ratio of association rate constants ( k ass ) of mAbs and bAbs was about two, due to the statistical factor of two binding sites per bivalent antibody molecule. However, the dissociation rate constant ( k diss ) of anti-HRP shoulder of bAbs was 21 times higher k diss of anti-HRP mAbs. The comparison with the theoretical model shows that these observations are consistent only with a situation in which bivalent binding of mAbs with Immobilized HRP predominates over monovalent binding. On the contrary, the second parental mAb (anti-hIgG) did not show the increase in avidity due to bivalent binding, compared to the anti-hIgG shoulder of bAbs, suggesting that this mAb was bound monovalently to Immobilized hIgG. The K ass values determined by solid-phase radioimmunoassay (RIA) yielded figures almost overlapping with those obtained by IAsys. The results of the comparison of bAbs and mAbs are discussed from the viewpoint of the use of bAbs in heterogeneous systems. On the other hand, these data demonstrate that real-time analysis of antibody binding parameters in IAsys biosensor is valuable for the selection of mAbs and bAbs with desired features, for different fields of application.
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the comparison of the ability of monoclonal antibodies directed to different proteins human igg human myoglobin and hrp and bispecific antibodies derived thereof to bind antigens Immobilized on a surface of a solid phase
Clinica Chimica Acta, 2001Co-Authors: D A Dmitriev, Yulia S Massino, O L Segal, M Smirnova, E V Pavlova, G I Kolyaskina, A P Osipov, A M Egorov, A D DmitrievAbstract:Abstract Background: Bindings of mouse monoclonal antibodies (mAbs) and affinity purified bispecific antibodies (bAbs), derived thereof, to antigens adsorbed on immunoplates have been compared, using ELISA and RIA methods. Methods: The analysed panel of antibodies included mAbs specific to human myoglobin (Mb), human IgG (hIgG) and horseradish peroxidase (HRP) and biologically produced bAbs with double specificity to Mb and HRP, and to hIgG and HRP. Results: The degree of difference between different mAbs and corresponding bAbs varied markedly from antibody to antibody, depending on whether the parental mAbs could bind Immobilized antigens bivalently. The observed equilibrium binding constant (Kobs) for anti-HRP mAbs was 21–38 times higher that of anti-HRP site of bAbs (anti-hIgG/HRP or anti-Mb/HRP, respectively), due to bivalent binding of mAbs. Anti-Mb mAbs also bound bivalently with Immobilized Mb. On the contrary, anti-hIgG mAbs bound monovalently with Immobilized hIgG in the same conditions. The avidity of anti-Mb/HRP bAbs increased, if both antigens were simultaneously adsorbed on a solid phase. Conclusions: The obtained data indicate that the use of bAbs in heterogeneous immunoassays instead of traditional mAb-enzyme conjugates hardly can provide the significant gain in assay performance if parental mAbs bind bivalently.
O L Segal - One of the best experts on this subject based on the ideXlab platform.
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kinetic analysis of interactions between bispecific monoclonal antibodies and Immobilized antigens using a resonant mirror biosensor
Journal of Immunological Methods, 2003Co-Authors: D A Dmitriev, Yulia S Massino, O L SegalAbstract:A resonant mirror biosensor (IAsys) protocol is described for the comparative kinetic analysis of the ability of monoclonal antibodies (Mabs) and bispecific antibodies (Babs) to bind Immobilized antigens. The protocol has been optimized and validated using the panel of affinity-purified antibodies, including two parental Mabs, one specific to human immunoglobulin G (hIgG) and another specific to horseradish peroxidase (HRP), and a Bab derived thereof by cell fusion (anti-hIgG/HRP Bab). The real-time kinetic analysis of antigen-antibody interactions using this protocol allows to demonstrate the differences in the avidity of bivalently binding Mabs and monovalent Babs. As shown in our previous study [J. Immunol. Methods 261 (2002) 103], the observed equilibrium association constants (Kass) determined by IAsys using this protocol yield figures almost overlapping with those obtained by solid-phase radioimmunoassay (RIA). The described protocol is suited for the investigation of the effects of valency on the binding properties of antibodies. It also may be applied for the selection of Mabs and Babs with desired features, for different fields of application.
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analysis of bispecific monoclonal antibody binding to Immobilized antigens using an optical biosensor
Biochemistry, 2002Co-Authors: D A Dmitriev, Yulia S Massino, O L Segal, M Smirnova, E V Pavlova, G I Kolyaskina, K G Gurevich, O V Gnedenko, Yuri D Ivanov, A I ArchakovAbstract:The interaction between two different monoclonal antibodies (Mabs) and their corresponding bispecific antibodies (Babs) with Immobilized antigens was investigated using an optical biosensor (IAsys). The analyzed panel of affinity-purified antibodies included two parental Mabs (one of which was specific to human IgG (hIgG), and another one to horseradish peroxidase (HRP)), as well as Babs derived thereof (anti-hIgG/HRP). Babs resulting from the fusion of parental hybridomas bear two antigen-binding sites toward two different antigens and thus may interact with Immobilized antigen through only one antigen-binding site (monovalently). Using an IAsys biosensor this study shows that the bivalent binding of Mabs predominates over the monovalent binding with Immobilized HRP, whereas anti-hIgG parental Mabs were bound monovalently to the Immobilized hIgG. The observed equilibrium association constant (K(ass)) values obtained in our last work [1] by solid-phase radioimmunoassay are consistent with those constants obtained by IAsys. The K(ass) of anti-HRP Mabs was about 50 times higher than that of anti-HRP shoulder of Babs. The dissociation rate constant (k(diss)) for anti-HRP shoulder of Babs was 21 times higher than k(diss) for anti-HRP Mabs. The comparison of the kinetic parameters for bivalent anti-HRP Mabs and Babs derived from anti-Mb/HRP and anti-hIgG/HRP, allowed to calculate that 95% of bound anti-HRP Mabs are bivalently linked with Immobilized HRP, whereas only 5% of bound anti-HRP Mabs are monovalently linked. In general, the data obtained indicate that Babs bearing an enzyme-binding site may not be efficiently used instead of traditional antibody-enzyme conjugates in the case of binding of bivalent Mabs.
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analysis of the binding of bispecific monoclonal antibodies with Immobilized antigens human igg and horseradish peroxidase using a resonant mirror biosensor
Journal of Immunological Methods, 2002Co-Authors: D A Dmitriev, Yulia S Massino, O L Segal, M Smirnova, E V Pavlova, G I Kolyaskina, K G Gurevich, O V Gnedenko, Yuri D Ivanov, A I ArchakovAbstract:Abstract The interaction between two monoclonal antibodies (mAbs) and their corresponding bispecific antibody (bAb) with Immobilized antigens has been examined using a resonant mirror biosensor (IAsys). BAbs were produced by cell fusion. The analysed panel of affinity-purified antibodies included two parental mAbs, one specific to human IgG (hIgG), and another specific to horseradish peroxidase (HRP), and a bAb derived thereof (anti-hIgG/HRP). The real-time analysis showed the drastic differences in the avidity of bivalent anti-HRP mAbs and anti-HRP shoulder of bAbs. Thus, the observed equilibrium association constant ( K ass ) of anti-HRP mAbs was about 50 times higher that of anti-HRP shoulder of bAbs. The ratio of association rate constants ( k ass ) of mAbs and bAbs was about two, due to the statistical factor of two binding sites per bivalent antibody molecule. However, the dissociation rate constant ( k diss ) of anti-HRP shoulder of bAbs was 21 times higher k diss of anti-HRP mAbs. The comparison with the theoretical model shows that these observations are consistent only with a situation in which bivalent binding of mAbs with Immobilized HRP predominates over monovalent binding. On the contrary, the second parental mAb (anti-hIgG) did not show the increase in avidity due to bivalent binding, compared to the anti-hIgG shoulder of bAbs, suggesting that this mAb was bound monovalently to Immobilized hIgG. The K ass values determined by solid-phase radioimmunoassay (RIA) yielded figures almost overlapping with those obtained by IAsys. The results of the comparison of bAbs and mAbs are discussed from the viewpoint of the use of bAbs in heterogeneous systems. On the other hand, these data demonstrate that real-time analysis of antibody binding parameters in IAsys biosensor is valuable for the selection of mAbs and bAbs with desired features, for different fields of application.
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the comparison of the ability of monoclonal antibodies directed to different proteins human igg human myoglobin and hrp and bispecific antibodies derived thereof to bind antigens Immobilized on a surface of a solid phase
Clinica Chimica Acta, 2001Co-Authors: D A Dmitriev, Yulia S Massino, O L Segal, M Smirnova, E V Pavlova, G I Kolyaskina, A P Osipov, A M Egorov, A D DmitrievAbstract:Abstract Background: Bindings of mouse monoclonal antibodies (mAbs) and affinity purified bispecific antibodies (bAbs), derived thereof, to antigens adsorbed on immunoplates have been compared, using ELISA and RIA methods. Methods: The analysed panel of antibodies included mAbs specific to human myoglobin (Mb), human IgG (hIgG) and horseradish peroxidase (HRP) and biologically produced bAbs with double specificity to Mb and HRP, and to hIgG and HRP. Results: The degree of difference between different mAbs and corresponding bAbs varied markedly from antibody to antibody, depending on whether the parental mAbs could bind Immobilized antigens bivalently. The observed equilibrium binding constant (Kobs) for anti-HRP mAbs was 21–38 times higher that of anti-HRP site of bAbs (anti-hIgG/HRP or anti-Mb/HRP, respectively), due to bivalent binding of mAbs. Anti-Mb mAbs also bound bivalently with Immobilized Mb. On the contrary, anti-hIgG mAbs bound monovalently with Immobilized hIgG in the same conditions. The avidity of anti-Mb/HRP bAbs increased, if both antigens were simultaneously adsorbed on a solid phase. Conclusions: The obtained data indicate that the use of bAbs in heterogeneous immunoassays instead of traditional mAb-enzyme conjugates hardly can provide the significant gain in assay performance if parental mAbs bind bivalently.
Christian Bourdillon - One of the best experts on this subject based on the ideXlab platform.
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activation and diffusion in the kinetics of adsorption and molecular recognition on surfaces enzyme amplified electrochemical approach to biorecognition dynamics illustrated by the binding of antibodies to Immobilized antigens
Journal of the American Chemical Society, 1999Co-Authors: Christian Bourdillon, Christophe Demaille, Jacques Moiroux, Jeanmichel SaveantAbstract:An electrochemical method is proposed for investigating the dynamics of recognition between a biomolecule and an Immobilized receptor. It involves redox labeling of the solute molecule and monitoring the binding by the electrochemical response of the electrode onto which the receptor is Immobilized. With large biomolecules, as, for example, antigens and antibodies, leading to small surface concentrations, simple redox labeling may prove insufficient to obtain detectable responses. Redox enzymes are then advantageously used as labels thanks to the signal amplification offered by their catalytic properties. The applicability of the method is illustrated by the reaction of an Immobilized monolayer of goat IgG antigen (or of one Fab fragment) with an antigoat antibody labeled with glucose oxidase. Particular care is taken to free the kinetic data from the effect of diffusion. The latter factor may interfere whatever the detection technique. A full account of the combination between recognition kinetics and di...
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activation and diffusion in the kinetics of adsorption and molecular recognition on surfaces enzyme amplified electrochemical approach to biorecognition dynamics illustrated by the binding of antibodies to Immobilized antigens
Journal of the American Chemical Society, 1999Co-Authors: Christian Bourdillon, Christophe Demaille, Jacques Moiroux, Jeanmichel SaveantAbstract:An electrochemical method is proposed for investigating the dynamics of recognition between a biomolecule and an Immobilized receptor. It involves redox labeling of the solute molecule and monitori...
Yulia S Massino - One of the best experts on this subject based on the ideXlab platform.
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kinetic analysis of interactions between bispecific monoclonal antibodies and Immobilized antigens using a resonant mirror biosensor
Journal of Immunological Methods, 2003Co-Authors: D A Dmitriev, Yulia S Massino, O L SegalAbstract:A resonant mirror biosensor (IAsys) protocol is described for the comparative kinetic analysis of the ability of monoclonal antibodies (Mabs) and bispecific antibodies (Babs) to bind Immobilized antigens. The protocol has been optimized and validated using the panel of affinity-purified antibodies, including two parental Mabs, one specific to human immunoglobulin G (hIgG) and another specific to horseradish peroxidase (HRP), and a Bab derived thereof by cell fusion (anti-hIgG/HRP Bab). The real-time kinetic analysis of antigen-antibody interactions using this protocol allows to demonstrate the differences in the avidity of bivalently binding Mabs and monovalent Babs. As shown in our previous study [J. Immunol. Methods 261 (2002) 103], the observed equilibrium association constants (Kass) determined by IAsys using this protocol yield figures almost overlapping with those obtained by solid-phase radioimmunoassay (RIA). The described protocol is suited for the investigation of the effects of valency on the binding properties of antibodies. It also may be applied for the selection of Mabs and Babs with desired features, for different fields of application.
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analysis of bispecific monoclonal antibody binding to Immobilized antigens using an optical biosensor
Biochemistry, 2002Co-Authors: D A Dmitriev, Yulia S Massino, O L Segal, M Smirnova, E V Pavlova, G I Kolyaskina, K G Gurevich, O V Gnedenko, Yuri D Ivanov, A I ArchakovAbstract:The interaction between two different monoclonal antibodies (Mabs) and their corresponding bispecific antibodies (Babs) with Immobilized antigens was investigated using an optical biosensor (IAsys). The analyzed panel of affinity-purified antibodies included two parental Mabs (one of which was specific to human IgG (hIgG), and another one to horseradish peroxidase (HRP)), as well as Babs derived thereof (anti-hIgG/HRP). Babs resulting from the fusion of parental hybridomas bear two antigen-binding sites toward two different antigens and thus may interact with Immobilized antigen through only one antigen-binding site (monovalently). Using an IAsys biosensor this study shows that the bivalent binding of Mabs predominates over the monovalent binding with Immobilized HRP, whereas anti-hIgG parental Mabs were bound monovalently to the Immobilized hIgG. The observed equilibrium association constant (K(ass)) values obtained in our last work [1] by solid-phase radioimmunoassay are consistent with those constants obtained by IAsys. The K(ass) of anti-HRP Mabs was about 50 times higher than that of anti-HRP shoulder of Babs. The dissociation rate constant (k(diss)) for anti-HRP shoulder of Babs was 21 times higher than k(diss) for anti-HRP Mabs. The comparison of the kinetic parameters for bivalent anti-HRP Mabs and Babs derived from anti-Mb/HRP and anti-hIgG/HRP, allowed to calculate that 95% of bound anti-HRP Mabs are bivalently linked with Immobilized HRP, whereas only 5% of bound anti-HRP Mabs are monovalently linked. In general, the data obtained indicate that Babs bearing an enzyme-binding site may not be efficiently used instead of traditional antibody-enzyme conjugates in the case of binding of bivalent Mabs.
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analysis of the binding of bispecific monoclonal antibodies with Immobilized antigens human igg and horseradish peroxidase using a resonant mirror biosensor
Journal of Immunological Methods, 2002Co-Authors: D A Dmitriev, Yulia S Massino, O L Segal, M Smirnova, E V Pavlova, G I Kolyaskina, K G Gurevich, O V Gnedenko, Yuri D Ivanov, A I ArchakovAbstract:Abstract The interaction between two monoclonal antibodies (mAbs) and their corresponding bispecific antibody (bAb) with Immobilized antigens has been examined using a resonant mirror biosensor (IAsys). BAbs were produced by cell fusion. The analysed panel of affinity-purified antibodies included two parental mAbs, one specific to human IgG (hIgG), and another specific to horseradish peroxidase (HRP), and a bAb derived thereof (anti-hIgG/HRP). The real-time analysis showed the drastic differences in the avidity of bivalent anti-HRP mAbs and anti-HRP shoulder of bAbs. Thus, the observed equilibrium association constant ( K ass ) of anti-HRP mAbs was about 50 times higher that of anti-HRP shoulder of bAbs. The ratio of association rate constants ( k ass ) of mAbs and bAbs was about two, due to the statistical factor of two binding sites per bivalent antibody molecule. However, the dissociation rate constant ( k diss ) of anti-HRP shoulder of bAbs was 21 times higher k diss of anti-HRP mAbs. The comparison with the theoretical model shows that these observations are consistent only with a situation in which bivalent binding of mAbs with Immobilized HRP predominates over monovalent binding. On the contrary, the second parental mAb (anti-hIgG) did not show the increase in avidity due to bivalent binding, compared to the anti-hIgG shoulder of bAbs, suggesting that this mAb was bound monovalently to Immobilized hIgG. The K ass values determined by solid-phase radioimmunoassay (RIA) yielded figures almost overlapping with those obtained by IAsys. The results of the comparison of bAbs and mAbs are discussed from the viewpoint of the use of bAbs in heterogeneous systems. On the other hand, these data demonstrate that real-time analysis of antibody binding parameters in IAsys biosensor is valuable for the selection of mAbs and bAbs with desired features, for different fields of application.
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the comparison of the ability of monoclonal antibodies directed to different proteins human igg human myoglobin and hrp and bispecific antibodies derived thereof to bind antigens Immobilized on a surface of a solid phase
Clinica Chimica Acta, 2001Co-Authors: D A Dmitriev, Yulia S Massino, O L Segal, M Smirnova, E V Pavlova, G I Kolyaskina, A P Osipov, A M Egorov, A D DmitrievAbstract:Abstract Background: Bindings of mouse monoclonal antibodies (mAbs) and affinity purified bispecific antibodies (bAbs), derived thereof, to antigens adsorbed on immunoplates have been compared, using ELISA and RIA methods. Methods: The analysed panel of antibodies included mAbs specific to human myoglobin (Mb), human IgG (hIgG) and horseradish peroxidase (HRP) and biologically produced bAbs with double specificity to Mb and HRP, and to hIgG and HRP. Results: The degree of difference between different mAbs and corresponding bAbs varied markedly from antibody to antibody, depending on whether the parental mAbs could bind Immobilized antigens bivalently. The observed equilibrium binding constant (Kobs) for anti-HRP mAbs was 21–38 times higher that of anti-HRP site of bAbs (anti-hIgG/HRP or anti-Mb/HRP, respectively), due to bivalent binding of mAbs. Anti-Mb mAbs also bound bivalently with Immobilized Mb. On the contrary, anti-hIgG mAbs bound monovalently with Immobilized hIgG in the same conditions. The avidity of anti-Mb/HRP bAbs increased, if both antigens were simultaneously adsorbed on a solid phase. Conclusions: The obtained data indicate that the use of bAbs in heterogeneous immunoassays instead of traditional mAb-enzyme conjugates hardly can provide the significant gain in assay performance if parental mAbs bind bivalently.
