Development of a highly sensitive chemiluminescent enzyme immunoassay for fragmented cytokeratin 18 using new antibodies

Abstract Fragmented cytokeratin 18 (fCK18) released from epithelial cells undergoing apoptosis is widely studied in various diseases. However, fCK18 measurement is not utilized in clinical practice due to imprecise disease-state cutoff values. Therefore, we set out to generate new monoclonal antibod...

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Autores principales:	Minori Yamada, Akiko Eguchi, Koji Okuno, Koji Sakaguchi, Tetsuji Yamaguchi
Formato:	article
Lenguaje:	EN
Publicado:	Nature Portfolio 2021
Materias:	Medicine R Science Q
Acceso en línea:	https://doaj.org/article/7efd161cd1164461a9c4b5bcb61a353a
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spelling	oai:doaj.org-article:7efd161cd1164461a9c4b5bcb61a353a2021-12-02T18:50:03ZDevelopment of a highly sensitive chemiluminescent enzyme immunoassay for fragmented cytokeratin 18 using new antibodies10.1038/s41598-021-97439-52045-2322https://doaj.org/article/7efd161cd1164461a9c4b5bcb61a353a2021-09-01T00:00:00Zhttps://doi.org/10.1038/s41598-021-97439-5https://doaj.org/toc/2045-2322Abstract Fragmented cytokeratin 18 (fCK18) released from epithelial cells undergoing apoptosis is widely studied in various diseases. However, fCK18 measurement is not utilized in clinical practice due to imprecise disease-state cutoff values. Therefore, we set out to generate new monoclonal antibodies (mAbs) and a recombinant fCK18 (rfCK18) calibrator in an effort to develop a highly sensitive chemiluminescent enzyme immunoassay (CLEIA). New capture mAb (K18-624) had a high binding ability compared to the current commercial antibody. New detection mAb (K18-328) recognized 323S-340G of CK18. A rfCK18 was expressed in the soluble fraction of E. coli when the N-terminal region (260 amino acid residues) of CK18 was truncated. Analysis of performance and measurement of human fCK18 were evaluated using K18-624 and K18-328 in a highly sensitive CLEIA. The coefficients of variation (CV) for within-run and between-day repeatability were below 10% and the recoveries were in the range of 15%. The detection sensitivity was 0.056 ng/mL. Serum fCK18 levels were significantly increased in non-alcoholic steatohepatitis (NASH) patients when compared to healthy individuals. Our new fCK18 mAbs showed high affinity and sensitivity. CLEIA using our new antibodies will be useful in measuring fCK18 in human blood thereby generating accurate clinical diagnoses of human liver diseases.Minori YamadaAkiko EguchiKoji OkunoKoji SakaguchiTetsuji YamaguchiNature PortfolioarticleMedicineRScienceQENScientific Reports, Vol 11, Iss 1, Pp 1-9 (2021)
institution	DOAJ
collection	DOAJ
language	EN
topic	Medicine R Science Q
spellingShingle	Medicine R Science Q Minori Yamada Akiko Eguchi Koji Okuno Koji Sakaguchi Tetsuji Yamaguchi Development of a highly sensitive chemiluminescent enzyme immunoassay for fragmented cytokeratin 18 using new antibodies
description	Abstract Fragmented cytokeratin 18 (fCK18) released from epithelial cells undergoing apoptosis is widely studied in various diseases. However, fCK18 measurement is not utilized in clinical practice due to imprecise disease-state cutoff values. Therefore, we set out to generate new monoclonal antibodies (mAbs) and a recombinant fCK18 (rfCK18) calibrator in an effort to develop a highly sensitive chemiluminescent enzyme immunoassay (CLEIA). New capture mAb (K18-624) had a high binding ability compared to the current commercial antibody. New detection mAb (K18-328) recognized 323S-340G of CK18. A rfCK18 was expressed in the soluble fraction of E. coli when the N-terminal region (260 amino acid residues) of CK18 was truncated. Analysis of performance and measurement of human fCK18 were evaluated using K18-624 and K18-328 in a highly sensitive CLEIA. The coefficients of variation (CV) for within-run and between-day repeatability were below 10% and the recoveries were in the range of 15%. The detection sensitivity was 0.056 ng/mL. Serum fCK18 levels were significantly increased in non-alcoholic steatohepatitis (NASH) patients when compared to healthy individuals. Our new fCK18 mAbs showed high affinity and sensitivity. CLEIA using our new antibodies will be useful in measuring fCK18 in human blood thereby generating accurate clinical diagnoses of human liver diseases.
format	article
author	Minori Yamada Akiko Eguchi Koji Okuno Koji Sakaguchi Tetsuji Yamaguchi
author_facet	Minori Yamada Akiko Eguchi Koji Okuno Koji Sakaguchi Tetsuji Yamaguchi
author_sort	Minori Yamada
title	Development of a highly sensitive chemiluminescent enzyme immunoassay for fragmented cytokeratin 18 using new antibodies
title_short	Development of a highly sensitive chemiluminescent enzyme immunoassay for fragmented cytokeratin 18 using new antibodies
title_full	Development of a highly sensitive chemiluminescent enzyme immunoassay for fragmented cytokeratin 18 using new antibodies
title_fullStr	Development of a highly sensitive chemiluminescent enzyme immunoassay for fragmented cytokeratin 18 using new antibodies
title_full_unstemmed	Development of a highly sensitive chemiluminescent enzyme immunoassay for fragmented cytokeratin 18 using new antibodies
title_sort	development of a highly sensitive chemiluminescent enzyme immunoassay for fragmented cytokeratin 18 using new antibodies
publisher	Nature Portfolio
publishDate	2021
url	https://doaj.org/article/7efd161cd1164461a9c4b5bcb61a353a
work_keys_str_mv	AT minoriyamada developmentofahighlysensitivechemiluminescentenzymeimmunoassayforfragmentedcytokeratin18usingnewantibodies AT akikoeguchi developmentofahighlysensitivechemiluminescentenzymeimmunoassayforfragmentedcytokeratin18usingnewantibodies AT kojiokuno developmentofahighlysensitivechemiluminescentenzymeimmunoassayforfragmentedcytokeratin18usingnewantibodies AT kojisakaguchi developmentofahighlysensitivechemiluminescentenzymeimmunoassayforfragmentedcytokeratin18usingnewantibodies AT tetsujiyamaguchi developmentofahighlysensitivechemiluminescentenzymeimmunoassayforfragmentedcytokeratin18usingnewantibodies
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Development of a highly sensitive chemiluminescent enzyme immunoassay for fragmented cytokeratin 18 using new antibodies

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