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Comparison of different chelates for lanthanide labeling of antibodies and application in a Western blot immunoassay combined with detection by laser ablation (LA-)ICP-MS

  • We have developed lanthanide labeling strategies for antibodies to adapt conventional biochemical workflows like Western blot immunoassays for detection by laser ablation inductively coupled plasma mass spectrometry (LA-ICP-MS) analysis with a special interest to apply the multi-element capabilities of ICP-MS for the design of multiplexed immunoassays. In this paper the lanthanide labeling of antibodies with MeCAT was investigated and the reaction conditions were optimized for application in a Western blot immunoassay analyzed by LA-ICP-MS. Furthermore, the MeCAT labeling strategy was compared with two other commercially available labeling reagents, MAXPAR™ and SCN-DOTA. As a proof-of-principle experiment chemically induced alterations of cytochrome P450 protein expression were investigated and the suitability of the differentially labeled antibodies for Western blot immunoassays of a complex liver microsomal protein fraction was tested. Limits of detection (LODs) in the lower fmolWe have developed lanthanide labeling strategies for antibodies to adapt conventional biochemical workflows like Western blot immunoassays for detection by laser ablation inductively coupled plasma mass spectrometry (LA-ICP-MS) analysis with a special interest to apply the multi-element capabilities of ICP-MS for the design of multiplexed immunoassays. In this paper the lanthanide labeling of antibodies with MeCAT was investigated and the reaction conditions were optimized for application in a Western blot immunoassay analyzed by LA-ICP-MS. Furthermore, the MeCAT labeling strategy was compared with two other commercially available labeling reagents, MAXPAR™ and SCN-DOTA. As a proof-of-principle experiment chemically induced alterations of cytochrome P450 protein expression were investigated and the suitability of the differentially labeled antibodies for Western blot immunoassays of a complex liver microsomal protein fraction was tested. Limits of detection (LODs) in the lower fmol range were reached in the Western blot application using MeCAT and MAXPAR™ as element labeling reagents, whereas even sub-fmol LODs can be achieved in a dot blot experiment for the pure antibodies.zeige mehrzeige weniger

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Metadaten
Autor*innen:Larissa Müller, Norbert JakubowskiORCiD, Simone Hösl, C. Scheler, P.H. Roos, M.W. Linscheid
Dokumenttyp:Zeitschriftenartikel
Veröffentlichungsform:Verlagsliteratur
Sprache:Englisch
Titel des übergeordneten Werkes (Englisch):Journal of analytical atomic spectrometry
Jahr der Erstveröffentlichung:2012
Verlag:Royal Society of Chemistry
Verlagsort:London
Jahrgang/Band:27
Ausgabe/Heft:8
Erste Seite:1311
Letzte Seite:1320
DOI:10.1039/c2ja30068k
ISSN:0267-9477
ISSN:1364-5544
Bemerkung:
Geburtsname von Müller, Larissa: Wäntig, L. -  Birth name of Müller, Larissa: Wäntig, L.
Bemerkung:
Geburtsname von Hösl, Simone: Hardt, S. -  Birth name of Hösl, Simone: Hardt, S.
Verfügbarkeit des Dokuments:Physisches Exemplar in der Bibliothek der BAM vorhanden ("Hardcopy Access")
Bibliotheksstandort:Sonderstandort: Publica-Schrank
Datum der Freischaltung:19.02.2016
Referierte Publikation:Ja
Datum der Eintragung als referierte Publikation:07.08.2012
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