Development and Validation of Approach for the Detection of Neutralizing Antibodies Against Insulin (Glargine) in Human Blood Plasma
Introduction. IImmunogenicity identification of therapeutic proteins, such as human insulin analogues, is one of the most relevant and significant area in medicine and pharmaceuticals. Determination the possibility of producing neutralizing antibodies to insulin reducing the therapeutic effect of th...
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| Autor*in: |
N. B. Abramenko [verfasserIn] P. I. Vnukova [verfasserIn] E. S. Golovina [verfasserIn] I. E. Makarenko [verfasserIn] A. A. Mosikian [verfasserIn] A. G. Nikiforova [verfasserIn] P. V. Gremyakova [verfasserIn] V. I. Kazey [verfasserIn] |
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| Format: |
E-Artikel |
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| Sprache: |
Russisch |
| Erschienen: |
2019 |
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| Schlagwörter: |
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| Übergeordnetes Werk: |
In: Разработка и регистрация лекарственных средств - LLC Center of Pharmaceutical Analytics (LLC «CPHA»), 2020, 8(2019), 3, Seite 70-78 |
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| Übergeordnetes Werk: |
volume:8 ; year:2019 ; number:3 ; pages:70-78 |
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| DOI / URN: |
10.33380/2305-2066-2019-8-3-70-78 |
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| Katalog-ID: |
DOAJ084732865 |
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| 520 | |a Introduction. IImmunogenicity identification of therapeutic proteins, such as human insulin analogues, is one of the most relevant and significant area in medicine and pharmaceuticals. Determination the possibility of producing neutralizing antibodies to insulin reducing the therapeutic effect of the drug, is an important step to understand the pharmacological profile of the drug. Applying of cell-based methods one allows to determinate neutralizing antibodies to insulin.Aim. Development and validation methods for detection of neutralizing antibodies against insulin in human plasma.Materials and methods. The method is based on the use of the iLiteTM Insulin Assay Ready Cells [1], in the genome of which the firefly luciferase reporter gene is introduced under the control of an insulin-dependent promoter. As the insulin concentration increases, the firefly luciferase expression (Firefly) increases, allowing one to use this cell line to estimate the number of neutralizing antibodies against insulin. For normalization by the number of cells and considering the matrix effect of studied samples, the second reporter gene luciferase Renilla is used, which is expressed under the control of a constitutive promoter. The activity of both luciferases was measured using the DualGlo Luciferase Assay System (Promega) assay [2].Results and discussion. Optimal insulin concentration and plasma/serum dilution were determined to identify neutralizing antibodies to insulin. The long-term stability of neutralizing antibodies to insulin were shown in human plasma for more than 3 months. The developed method was applied in a comparative research of the safety and immunogenicity of insulin analogues (Glargine). Method for the determination of antibodies to insulin was.Conclusion. A method for determination of neutralizing antibodies to insulin in human K2EDTA plasma was developed and validated using iLiteTM Insulin Assay Ready Cells system; based on the binding of the insulin alpha chain to the high-affinity heterodimeric CD220 receptor. | ||
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10.33380/2305-2066-2019-8-3-70-78 doi (DE-627)DOAJ084732865 (DE-599)DOAJ9e0df99d461c4dada32752a690fe9619 DE-627 ger DE-627 rakwb rus HD9665-9675 N. B. Abramenko verfasserin aut Development and Validation of Approach for the Detection of Neutralizing Antibodies Against Insulin (Glargine) in Human Blood Plasma 2019 Text txt rdacontent Computermedien c rdamedia Online-Ressource cr rdacarrier Introduction. IImmunogenicity identification of therapeutic proteins, such as human insulin analogues, is one of the most relevant and significant area in medicine and pharmaceuticals. Determination the possibility of producing neutralizing antibodies to insulin reducing the therapeutic effect of the drug, is an important step to understand the pharmacological profile of the drug. Applying of cell-based methods one allows to determinate neutralizing antibodies to insulin.Aim. Development and validation methods for detection of neutralizing antibodies against insulin in human plasma.Materials and methods. The method is based on the use of the iLiteTM Insulin Assay Ready Cells [1], in the genome of which the firefly luciferase reporter gene is introduced under the control of an insulin-dependent promoter. As the insulin concentration increases, the firefly luciferase expression (Firefly) increases, allowing one to use this cell line to estimate the number of neutralizing antibodies against insulin. For normalization by the number of cells and considering the matrix effect of studied samples, the second reporter gene luciferase Renilla is used, which is expressed under the control of a constitutive promoter. The activity of both luciferases was measured using the DualGlo Luciferase Assay System (Promega) assay [2].Results and discussion. Optimal insulin concentration and plasma/serum dilution were determined to identify neutralizing antibodies to insulin. The long-term stability of neutralizing antibodies to insulin were shown in human plasma for more than 3 months. The developed method was applied in a comparative research of the safety and immunogenicity of insulin analogues (Glargine). Method for the determination of antibodies to insulin was.Conclusion. A method for determination of neutralizing antibodies to insulin in human K2EDTA plasma was developed and validated using iLiteTM Insulin Assay Ready Cells system; based on the binding of the insulin alpha chain to the high-affinity heterodimeric CD220 receptor. нейтрализующие антитела инсулин гларгин клеточные методы валидация Pharmaceutical industry P. I. Vnukova verfasserin aut E. S. Golovina verfasserin aut I. E. Makarenko verfasserin aut A. A. Mosikian verfasserin aut A. G. Nikiforova verfasserin aut P. V. Gremyakova verfasserin aut V. I. Kazey verfasserin aut In Разработка и регистрация лекарственных средств LLC Center of Pharmaceutical Analytics (LLC «CPHA»), 2020 8(2019), 3, Seite 70-78 (DE-627)1760622966 26585049 nnns volume:8 year:2019 number:3 pages:70-78 https://doi.org/10.33380/2305-2066-2019-8-3-70-78 kostenfrei https://doaj.org/article/9e0df99d461c4dada32752a690fe9619 kostenfrei https://www.pharmjournal.ru/jour/article/view/717 kostenfrei https://doaj.org/toc/2305-2066 Journal toc kostenfrei https://doaj.org/toc/2658-5049 Journal toc kostenfrei GBV_USEFLAG_A SYSFLAG_A GBV_DOAJ AR 8 2019 3 70-78 |
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10.33380/2305-2066-2019-8-3-70-78 doi (DE-627)DOAJ084732865 (DE-599)DOAJ9e0df99d461c4dada32752a690fe9619 DE-627 ger DE-627 rakwb rus HD9665-9675 N. B. Abramenko verfasserin aut Development and Validation of Approach for the Detection of Neutralizing Antibodies Against Insulin (Glargine) in Human Blood Plasma 2019 Text txt rdacontent Computermedien c rdamedia Online-Ressource cr rdacarrier Introduction. IImmunogenicity identification of therapeutic proteins, such as human insulin analogues, is one of the most relevant and significant area in medicine and pharmaceuticals. Determination the possibility of producing neutralizing antibodies to insulin reducing the therapeutic effect of the drug, is an important step to understand the pharmacological profile of the drug. Applying of cell-based methods one allows to determinate neutralizing antibodies to insulin.Aim. Development and validation methods for detection of neutralizing antibodies against insulin in human plasma.Materials and methods. The method is based on the use of the iLiteTM Insulin Assay Ready Cells [1], in the genome of which the firefly luciferase reporter gene is introduced under the control of an insulin-dependent promoter. As the insulin concentration increases, the firefly luciferase expression (Firefly) increases, allowing one to use this cell line to estimate the number of neutralizing antibodies against insulin. For normalization by the number of cells and considering the matrix effect of studied samples, the second reporter gene luciferase Renilla is used, which is expressed under the control of a constitutive promoter. The activity of both luciferases was measured using the DualGlo Luciferase Assay System (Promega) assay [2].Results and discussion. Optimal insulin concentration and plasma/serum dilution were determined to identify neutralizing antibodies to insulin. The long-term stability of neutralizing antibodies to insulin were shown in human plasma for more than 3 months. The developed method was applied in a comparative research of the safety and immunogenicity of insulin analogues (Glargine). Method for the determination of antibodies to insulin was.Conclusion. A method for determination of neutralizing antibodies to insulin in human K2EDTA plasma was developed and validated using iLiteTM Insulin Assay Ready Cells system; based on the binding of the insulin alpha chain to the high-affinity heterodimeric CD220 receptor. нейтрализующие антитела инсулин гларгин клеточные методы валидация Pharmaceutical industry P. I. Vnukova verfasserin aut E. S. Golovina verfasserin aut I. E. Makarenko verfasserin aut A. A. Mosikian verfasserin aut A. G. Nikiforova verfasserin aut P. V. Gremyakova verfasserin aut V. I. Kazey verfasserin aut In Разработка и регистрация лекарственных средств LLC Center of Pharmaceutical Analytics (LLC «CPHA»), 2020 8(2019), 3, Seite 70-78 (DE-627)1760622966 26585049 nnns volume:8 year:2019 number:3 pages:70-78 https://doi.org/10.33380/2305-2066-2019-8-3-70-78 kostenfrei https://doaj.org/article/9e0df99d461c4dada32752a690fe9619 kostenfrei https://www.pharmjournal.ru/jour/article/view/717 kostenfrei https://doaj.org/toc/2305-2066 Journal toc kostenfrei https://doaj.org/toc/2658-5049 Journal toc kostenfrei GBV_USEFLAG_A SYSFLAG_A GBV_DOAJ AR 8 2019 3 70-78 |
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10.33380/2305-2066-2019-8-3-70-78 doi (DE-627)DOAJ084732865 (DE-599)DOAJ9e0df99d461c4dada32752a690fe9619 DE-627 ger DE-627 rakwb rus HD9665-9675 N. B. Abramenko verfasserin aut Development and Validation of Approach for the Detection of Neutralizing Antibodies Against Insulin (Glargine) in Human Blood Plasma 2019 Text txt rdacontent Computermedien c rdamedia Online-Ressource cr rdacarrier Introduction. IImmunogenicity identification of therapeutic proteins, such as human insulin analogues, is one of the most relevant and significant area in medicine and pharmaceuticals. Determination the possibility of producing neutralizing antibodies to insulin reducing the therapeutic effect of the drug, is an important step to understand the pharmacological profile of the drug. Applying of cell-based methods one allows to determinate neutralizing antibodies to insulin.Aim. Development and validation methods for detection of neutralizing antibodies against insulin in human plasma.Materials and methods. The method is based on the use of the iLiteTM Insulin Assay Ready Cells [1], in the genome of which the firefly luciferase reporter gene is introduced under the control of an insulin-dependent promoter. As the insulin concentration increases, the firefly luciferase expression (Firefly) increases, allowing one to use this cell line to estimate the number of neutralizing antibodies against insulin. For normalization by the number of cells and considering the matrix effect of studied samples, the second reporter gene luciferase Renilla is used, which is expressed under the control of a constitutive promoter. The activity of both luciferases was measured using the DualGlo Luciferase Assay System (Promega) assay [2].Results and discussion. Optimal insulin concentration and plasma/serum dilution were determined to identify neutralizing antibodies to insulin. The long-term stability of neutralizing antibodies to insulin were shown in human plasma for more than 3 months. The developed method was applied in a comparative research of the safety and immunogenicity of insulin analogues (Glargine). Method for the determination of antibodies to insulin was.Conclusion. A method for determination of neutralizing antibodies to insulin in human K2EDTA plasma was developed and validated using iLiteTM Insulin Assay Ready Cells system; based on the binding of the insulin alpha chain to the high-affinity heterodimeric CD220 receptor. нейтрализующие антитела инсулин гларгин клеточные методы валидация Pharmaceutical industry P. I. Vnukova verfasserin aut E. S. Golovina verfasserin aut I. E. Makarenko verfasserin aut A. A. Mosikian verfasserin aut A. G. Nikiforova verfasserin aut P. V. Gremyakova verfasserin aut V. I. Kazey verfasserin aut In Разработка и регистрация лекарственных средств LLC Center of Pharmaceutical Analytics (LLC «CPHA»), 2020 8(2019), 3, Seite 70-78 (DE-627)1760622966 26585049 nnns volume:8 year:2019 number:3 pages:70-78 https://doi.org/10.33380/2305-2066-2019-8-3-70-78 kostenfrei https://doaj.org/article/9e0df99d461c4dada32752a690fe9619 kostenfrei https://www.pharmjournal.ru/jour/article/view/717 kostenfrei https://doaj.org/toc/2305-2066 Journal toc kostenfrei https://doaj.org/toc/2658-5049 Journal toc kostenfrei GBV_USEFLAG_A SYSFLAG_A GBV_DOAJ AR 8 2019 3 70-78 |
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10.33380/2305-2066-2019-8-3-70-78 doi (DE-627)DOAJ084732865 (DE-599)DOAJ9e0df99d461c4dada32752a690fe9619 DE-627 ger DE-627 rakwb rus HD9665-9675 N. B. Abramenko verfasserin aut Development and Validation of Approach for the Detection of Neutralizing Antibodies Against Insulin (Glargine) in Human Blood Plasma 2019 Text txt rdacontent Computermedien c rdamedia Online-Ressource cr rdacarrier Introduction. IImmunogenicity identification of therapeutic proteins, such as human insulin analogues, is one of the most relevant and significant area in medicine and pharmaceuticals. Determination the possibility of producing neutralizing antibodies to insulin reducing the therapeutic effect of the drug, is an important step to understand the pharmacological profile of the drug. Applying of cell-based methods one allows to determinate neutralizing antibodies to insulin.Aim. Development and validation methods for detection of neutralizing antibodies against insulin in human plasma.Materials and methods. The method is based on the use of the iLiteTM Insulin Assay Ready Cells [1], in the genome of which the firefly luciferase reporter gene is introduced under the control of an insulin-dependent promoter. As the insulin concentration increases, the firefly luciferase expression (Firefly) increases, allowing one to use this cell line to estimate the number of neutralizing antibodies against insulin. For normalization by the number of cells and considering the matrix effect of studied samples, the second reporter gene luciferase Renilla is used, which is expressed under the control of a constitutive promoter. The activity of both luciferases was measured using the DualGlo Luciferase Assay System (Promega) assay [2].Results and discussion. Optimal insulin concentration and plasma/serum dilution were determined to identify neutralizing antibodies to insulin. The long-term stability of neutralizing antibodies to insulin were shown in human plasma for more than 3 months. The developed method was applied in a comparative research of the safety and immunogenicity of insulin analogues (Glargine). Method for the determination of antibodies to insulin was.Conclusion. A method for determination of neutralizing antibodies to insulin in human K2EDTA plasma was developed and validated using iLiteTM Insulin Assay Ready Cells system; based on the binding of the insulin alpha chain to the high-affinity heterodimeric CD220 receptor. нейтрализующие антитела инсулин гларгин клеточные методы валидация Pharmaceutical industry P. I. Vnukova verfasserin aut E. S. Golovina verfasserin aut I. E. Makarenko verfasserin aut A. A. Mosikian verfasserin aut A. G. Nikiforova verfasserin aut P. V. Gremyakova verfasserin aut V. I. Kazey verfasserin aut In Разработка и регистрация лекарственных средств LLC Center of Pharmaceutical Analytics (LLC «CPHA»), 2020 8(2019), 3, Seite 70-78 (DE-627)1760622966 26585049 nnns volume:8 year:2019 number:3 pages:70-78 https://doi.org/10.33380/2305-2066-2019-8-3-70-78 kostenfrei https://doaj.org/article/9e0df99d461c4dada32752a690fe9619 kostenfrei https://www.pharmjournal.ru/jour/article/view/717 kostenfrei https://doaj.org/toc/2305-2066 Journal toc kostenfrei https://doaj.org/toc/2658-5049 Journal toc kostenfrei GBV_USEFLAG_A SYSFLAG_A GBV_DOAJ AR 8 2019 3 70-78 |
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10.33380/2305-2066-2019-8-3-70-78 doi (DE-627)DOAJ084732865 (DE-599)DOAJ9e0df99d461c4dada32752a690fe9619 DE-627 ger DE-627 rakwb rus HD9665-9675 N. B. Abramenko verfasserin aut Development and Validation of Approach for the Detection of Neutralizing Antibodies Against Insulin (Glargine) in Human Blood Plasma 2019 Text txt rdacontent Computermedien c rdamedia Online-Ressource cr rdacarrier Introduction. IImmunogenicity identification of therapeutic proteins, such as human insulin analogues, is one of the most relevant and significant area in medicine and pharmaceuticals. Determination the possibility of producing neutralizing antibodies to insulin reducing the therapeutic effect of the drug, is an important step to understand the pharmacological profile of the drug. Applying of cell-based methods one allows to determinate neutralizing antibodies to insulin.Aim. Development and validation methods for detection of neutralizing antibodies against insulin in human plasma.Materials and methods. The method is based on the use of the iLiteTM Insulin Assay Ready Cells [1], in the genome of which the firefly luciferase reporter gene is introduced under the control of an insulin-dependent promoter. As the insulin concentration increases, the firefly luciferase expression (Firefly) increases, allowing one to use this cell line to estimate the number of neutralizing antibodies against insulin. For normalization by the number of cells and considering the matrix effect of studied samples, the second reporter gene luciferase Renilla is used, which is expressed under the control of a constitutive promoter. The activity of both luciferases was measured using the DualGlo Luciferase Assay System (Promega) assay [2].Results and discussion. Optimal insulin concentration and plasma/serum dilution were determined to identify neutralizing antibodies to insulin. The long-term stability of neutralizing antibodies to insulin were shown in human plasma for more than 3 months. The developed method was applied in a comparative research of the safety and immunogenicity of insulin analogues (Glargine). Method for the determination of antibodies to insulin was.Conclusion. A method for determination of neutralizing antibodies to insulin in human K2EDTA plasma was developed and validated using iLiteTM Insulin Assay Ready Cells system; based on the binding of the insulin alpha chain to the high-affinity heterodimeric CD220 receptor. нейтрализующие антитела инсулин гларгин клеточные методы валидация Pharmaceutical industry P. I. Vnukova verfasserin aut E. S. Golovina verfasserin aut I. E. Makarenko verfasserin aut A. A. Mosikian verfasserin aut A. G. Nikiforova verfasserin aut P. V. Gremyakova verfasserin aut V. I. Kazey verfasserin aut In Разработка и регистрация лекарственных средств LLC Center of Pharmaceutical Analytics (LLC «CPHA»), 2020 8(2019), 3, Seite 70-78 (DE-627)1760622966 26585049 nnns volume:8 year:2019 number:3 pages:70-78 https://doi.org/10.33380/2305-2066-2019-8-3-70-78 kostenfrei https://doaj.org/article/9e0df99d461c4dada32752a690fe9619 kostenfrei https://www.pharmjournal.ru/jour/article/view/717 kostenfrei https://doaj.org/toc/2305-2066 Journal toc kostenfrei https://doaj.org/toc/2658-5049 Journal toc kostenfrei GBV_USEFLAG_A SYSFLAG_A GBV_DOAJ AR 8 2019 3 70-78 |
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B. Abramenko</subfield><subfield code="e">verfasserin</subfield><subfield code="4">aut</subfield></datafield><datafield tag="245" ind1="1" ind2="0"><subfield code="a">Development and Validation of Approach for the Detection of Neutralizing Antibodies Against Insulin (Glargine) in Human Blood Plasma</subfield></datafield><datafield tag="264" ind1=" " ind2="1"><subfield code="c">2019</subfield></datafield><datafield tag="336" ind1=" " ind2=" "><subfield code="a">Text</subfield><subfield code="b">txt</subfield><subfield code="2">rdacontent</subfield></datafield><datafield tag="337" ind1=" " ind2=" "><subfield code="a">Computermedien</subfield><subfield code="b">c</subfield><subfield code="2">rdamedia</subfield></datafield><datafield tag="338" ind1=" " ind2=" "><subfield code="a">Online-Ressource</subfield><subfield code="b">cr</subfield><subfield code="2">rdacarrier</subfield></datafield><datafield tag="520" ind1=" " ind2=" "><subfield code="a">Introduction. IImmunogenicity identification of therapeutic proteins, such as human insulin analogues, is one of the most relevant and significant area in medicine and pharmaceuticals. Determination the possibility of producing neutralizing antibodies to insulin reducing the therapeutic effect of the drug, is an important step to understand the pharmacological profile of the drug. Applying of cell-based methods one allows to determinate neutralizing antibodies to insulin.Aim. Development and validation methods for detection of neutralizing antibodies against insulin in human plasma.Materials and methods. The method is based on the use of the iLiteTM Insulin Assay Ready Cells [1], in the genome of which the firefly luciferase reporter gene is introduced under the control of an insulin-dependent promoter. As the insulin concentration increases, the firefly luciferase expression (Firefly) increases, allowing one to use this cell line to estimate the number of neutralizing antibodies against insulin. For normalization by the number of cells and considering the matrix effect of studied samples, the second reporter gene luciferase Renilla is used, which is expressed under the control of a constitutive promoter. The activity of both luciferases was measured using the DualGlo Luciferase Assay System (Promega) assay [2].Results and discussion. Optimal insulin concentration and plasma/serum dilution were determined to identify neutralizing antibodies to insulin. The long-term stability of neutralizing antibodies to insulin were shown in human plasma for more than 3 months. The developed method was applied in a comparative research of the safety and immunogenicity of insulin analogues (Glargine). Method for the determination of antibodies to insulin was.Conclusion. 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HD9665-9675 Development and Validation of Approach for the Detection of Neutralizing Antibodies Against Insulin (Glargine) in Human Blood Plasma нейтрализующие антитела инсулин гларгин клеточные методы валидация |
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N. B. Abramenko P. I. Vnukova E. S. Golovina I. E. Makarenko A. A. Mosikian A. G. Nikiforova P. V. Gremyakova V. I. Kazey |
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development and validation of approach for the detection of neutralizing antibodies against insulin (glargine) in human blood plasma |
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HD9665-9675 |
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Development and Validation of Approach for the Detection of Neutralizing Antibodies Against Insulin (Glargine) in Human Blood Plasma |
| abstract |
Introduction. IImmunogenicity identification of therapeutic proteins, such as human insulin analogues, is one of the most relevant and significant area in medicine and pharmaceuticals. Determination the possibility of producing neutralizing antibodies to insulin reducing the therapeutic effect of the drug, is an important step to understand the pharmacological profile of the drug. Applying of cell-based methods one allows to determinate neutralizing antibodies to insulin.Aim. Development and validation methods for detection of neutralizing antibodies against insulin in human plasma.Materials and methods. The method is based on the use of the iLiteTM Insulin Assay Ready Cells [1], in the genome of which the firefly luciferase reporter gene is introduced under the control of an insulin-dependent promoter. As the insulin concentration increases, the firefly luciferase expression (Firefly) increases, allowing one to use this cell line to estimate the number of neutralizing antibodies against insulin. For normalization by the number of cells and considering the matrix effect of studied samples, the second reporter gene luciferase Renilla is used, which is expressed under the control of a constitutive promoter. The activity of both luciferases was measured using the DualGlo Luciferase Assay System (Promega) assay [2].Results and discussion. Optimal insulin concentration and plasma/serum dilution were determined to identify neutralizing antibodies to insulin. The long-term stability of neutralizing antibodies to insulin were shown in human plasma for more than 3 months. The developed method was applied in a comparative research of the safety and immunogenicity of insulin analogues (Glargine). Method for the determination of antibodies to insulin was.Conclusion. A method for determination of neutralizing antibodies to insulin in human K2EDTA plasma was developed and validated using iLiteTM Insulin Assay Ready Cells system; based on the binding of the insulin alpha chain to the high-affinity heterodimeric CD220 receptor. |
| abstractGer |
Introduction. IImmunogenicity identification of therapeutic proteins, such as human insulin analogues, is one of the most relevant and significant area in medicine and pharmaceuticals. Determination the possibility of producing neutralizing antibodies to insulin reducing the therapeutic effect of the drug, is an important step to understand the pharmacological profile of the drug. Applying of cell-based methods one allows to determinate neutralizing antibodies to insulin.Aim. Development and validation methods for detection of neutralizing antibodies against insulin in human plasma.Materials and methods. The method is based on the use of the iLiteTM Insulin Assay Ready Cells [1], in the genome of which the firefly luciferase reporter gene is introduced under the control of an insulin-dependent promoter. As the insulin concentration increases, the firefly luciferase expression (Firefly) increases, allowing one to use this cell line to estimate the number of neutralizing antibodies against insulin. For normalization by the number of cells and considering the matrix effect of studied samples, the second reporter gene luciferase Renilla is used, which is expressed under the control of a constitutive promoter. The activity of both luciferases was measured using the DualGlo Luciferase Assay System (Promega) assay [2].Results and discussion. Optimal insulin concentration and plasma/serum dilution were determined to identify neutralizing antibodies to insulin. The long-term stability of neutralizing antibodies to insulin were shown in human plasma for more than 3 months. The developed method was applied in a comparative research of the safety and immunogenicity of insulin analogues (Glargine). Method for the determination of antibodies to insulin was.Conclusion. A method for determination of neutralizing antibodies to insulin in human K2EDTA plasma was developed and validated using iLiteTM Insulin Assay Ready Cells system; based on the binding of the insulin alpha chain to the high-affinity heterodimeric CD220 receptor. |
| abstract_unstemmed |
Introduction. IImmunogenicity identification of therapeutic proteins, such as human insulin analogues, is one of the most relevant and significant area in medicine and pharmaceuticals. Determination the possibility of producing neutralizing antibodies to insulin reducing the therapeutic effect of the drug, is an important step to understand the pharmacological profile of the drug. Applying of cell-based methods one allows to determinate neutralizing antibodies to insulin.Aim. Development and validation methods for detection of neutralizing antibodies against insulin in human plasma.Materials and methods. The method is based on the use of the iLiteTM Insulin Assay Ready Cells [1], in the genome of which the firefly luciferase reporter gene is introduced under the control of an insulin-dependent promoter. As the insulin concentration increases, the firefly luciferase expression (Firefly) increases, allowing one to use this cell line to estimate the number of neutralizing antibodies against insulin. For normalization by the number of cells and considering the matrix effect of studied samples, the second reporter gene luciferase Renilla is used, which is expressed under the control of a constitutive promoter. The activity of both luciferases was measured using the DualGlo Luciferase Assay System (Promega) assay [2].Results and discussion. Optimal insulin concentration and plasma/serum dilution were determined to identify neutralizing antibodies to insulin. The long-term stability of neutralizing antibodies to insulin were shown in human plasma for more than 3 months. The developed method was applied in a comparative research of the safety and immunogenicity of insulin analogues (Glargine). Method for the determination of antibodies to insulin was.Conclusion. A method for determination of neutralizing antibodies to insulin in human K2EDTA plasma was developed and validated using iLiteTM Insulin Assay Ready Cells system; based on the binding of the insulin alpha chain to the high-affinity heterodimeric CD220 receptor. |
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