A Novel Method to Assay Proteins in Blood Plasma after Intravenous Injection of Plasmid DNA

Hanawa, Haruo; Watanabe, Ritsuo; Hayashi, Manabu; Yoshida, Tsuyoshi; Abe, Satoru; Komura, Satoru; Liu, Hui; Elnaggar, Raafat; Chang, He; Okura, Yuji; Kato, Kiminori; Kodama, Makoto; Maruyama, Hiroki; Miyazaki, Junichi; Aizawa, Yoshifusa

doi:info:doi/10.1620/tjem.202.155

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タイトル	A Novel Method to Assay Proteins in Blood Plasma after Intravenous Injection of Plasmid DNA
著者	Hanawa, Haruo Hanawa, Haruo
	Watanabe, Ritsuo Watanabe, Ritsuo
	Hayashi, Manabu Hayashi, Manabu
	Yoshida, Tsuyoshi Yoshida, Tsuyoshi
	Abe, Satoru Abe, Satoru
	Komura, Satoru Komura, Satoru
	Liu, Hui Liu, Hui
	Elnaggar, Raafat Elnaggar, Raafat
	Chang, He Chang, He
	Okura, Yuji Okura, Yuji
	Kato, Kiminori Kato, Kiminori
	Kodama, Makoto Kodama, Makoto
	Maruyama, Hiroki Maruyama, Hiroki
	Miyazaki, Junichi Miyazaki, Junichi
	Aizawa, Yoshifusa Aizawa, Yoshifusa
抄録	Gene therapy is expected to lead to new and useful methods to treat diseases. The development of assays to quantitate gene-therapy-derived proteins circulating in blood will be essential to investigate the effects and side effects of the introduced proteins. The purpose of this study is to evaluate whether a protein circulating at trace concentrations in blood can be measured by tagging a peptide corresponding to glucagon residues 19-29 onto its C-terminal end. We constructed plasmids encoding chimeric proteins and transferred them into rats by hydrodynamics-based delivery. When plasmids encoding human IL8-glucagon 19-29 chimeric protein were injected into rats to evaluate the accuracy of this method, there was a high correlation between chimeric proteins measured by an enzyme-linked immunosorbent assay for human IL8 and one by a radioimmunoassay for glucagon. Furthermore, when plasmids coding rat IFN gamma receptor IgG-Fc glucagon 19-29 chimeric protein were injected to evaluate the time course of chimeric proteins in blood plasma, we could calculate the concentrations in blood from 10 μl plasma samples using glucagon 19-29 tag as follows: 2815±2318 ng/ml after 4 hours (mean±S.D.), 6061±2789 ng/ml after 8 hours, 5752±2270 ng/ml after 12 hours, 2870±1062 ng/ml after one day, 1440±334 ng/ml after three days, 1120±433 ng/ml after seven days, and 281±162 ng/ml after 16 days. Blood sugar levels which might have been increased by glucagon did not increase even at peak chimeric protein concentrations. These results demonstrate a useful and convenient method to assay gene therapy products circulating in blood using a glucagon 19-29 tagging vector.
公開者	Tohoku University Medical Press
公開者の別表記	東北ジャーナル刊行会
公開者 (ヨミ)	トウホク　ジャーナル　カンコウカイ
掲載誌名	Tohoku Journal of Experimental Medicine
巻	202
号	3
開始ページ	155
終了ページ	161
刊行年月	2004-01-09
ISSN	00408727
NCID	AA00863920
DOI	info:doi/10.1620/tjem.202.155
URL	http://hdl.handle.net/11094/23114
権利情報	© 2004 Tohoku University Medical Press
言語	英語
カテゴリ	学術雑誌論文 Journal Article

著者版フラグ	publisher
NII資源タイプ	学術雑誌論文
ローカル資源タイプ	学術雑誌論文
dcmi資源タイプ	text
DCTERMS.bibliographicCitation	Tohoku Journal of Experimental Medicine.202(3) P.155-P.161
DC.title	A Novel Method to Assay Proteins in Blood Plasma after Intravenous Injection of Plasmid DNA
DC.creator	Hanawa, Haruo
	Watanabe, Ritsuo
	Hayashi, Manabu
	Yoshida, Tsuyoshi
	Abe, Satoru
	Komura, Satoru
	Liu, Hui
	Elnaggar, Raafat
	Chang, He
	Okura, Yuji
	Kato, Kiminori
	Kodama, Makoto
	Maruyama, Hiroki
	Miyazaki, Junichi
	Aizawa, Yoshifusa
DC.publisher	Tohoku University Medical Press
DC.language" scheme="DCTERMS.RFC1766	英語
DCTERMS.issued" scheme="DCTERMS.W3CDTF	2004-01-09
DC.identifier	info:doi/10.1620/tjem.202.155
DC.identifier" scheme="DCTERMS.URI	http://hdl.handle.net/11094/23114
DCTERMS.abstract	Gene therapy is expected to lead to new and useful methods to treat diseases. The development of assays to quantitate gene-therapy-derived proteins circulating in blood will be essential to investigate the effects and side effects of the introduced proteins. The purpose of this study is to evaluate whether a protein circulating at trace concentrations in blood can be measured by tagging a peptide corresponding to glucagon residues 19-29 onto its C-terminal end. We constructed plasmids encoding chimeric proteins and transferred them into rats by hydrodynamics-based delivery. When plasmids encoding human IL8-glucagon 19-29 chimeric protein were injected into rats to evaluate the accuracy of this method, there was a high correlation between chimeric proteins measured by an enzyme-linked immunosorbent assay for human IL8 and one by a radioimmunoassay for glucagon. Furthermore, when plasmids coding rat IFN gamma receptor IgG-Fc glucagon 19-29 chimeric protein were injected to evaluate the time course of chimeric proteins in blood plasma, we could calculate the concentrations in blood from 10 μl plasma samples using glucagon 19-29 tag as follows: 2815±2318 ng/ml after 4 hours (mean±S.D.), 6061±2789 ng/ml after 8 hours, 5752±2270 ng/ml after 12 hours, 2870±1062 ng/ml after one day, 1440±334 ng/ml after three days, 1120±433 ng/ml after seven days, and 281±162 ng/ml after 16 days. Blood sugar levels which might have been increased by glucagon did not increase even at peak chimeric protein concentrations. These results demonstrate a useful and convenient method to assay gene therapy products circulating in blood using a glucagon 19-29 tagging vector.
DC.rights	© 2004 Tohoku University Medical Press
citation_title	A Novel Method to Assay Proteins in Blood Plasma after Intravenous Injection of Plasmid DNA
citation_author	Hanawa, Haruo
	Watanabe, Ritsuo
	Hayashi, Manabu
	Yoshida, Tsuyoshi
	Abe, Satoru
	Komura, Satoru
	Liu, Hui
	Elnaggar, Raafat
	Chang, He
	Okura, Yuji
	Kato, Kiminori
	Kodama, Makoto
	Maruyama, Hiroki
	Miyazaki, Junichi
	Aizawa, Yoshifusa
citation_publisher	Tohoku University Medical Press
citation_language	英語
citation_date	2004-01-09
citation_journal_title	Tohoku Journal of Experimental Medicine
citation_volume	202
citation_issue	3
citation_firstpage	155
citation_lastpage	161
citation_issn	00408727
citation_doi	info:doi/10.1620/tjem.202.155
citation_public_url	http://hdl.handle.net/11094/23114

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