Type

Journal Article

Authors

Lorraine O'Driscoll
Martin Clynes
Paul Barham
Finbarr O'Sullivan
Niall W Barron
Eadaoin Mc Kiernan

Subjects

Biochemistry

Topics
insulin tissue culture techniques animals metabolism embryonic stem cells c peptide cell lineage cells cultured biosynthesis mice biological markers cell differentiation cell line tumor dose response relationship drug physiology

Detecting de novo insulin synthesis in embryonic stem cell-derived populations. (2006)

Abstract Several studies in recent years have described protocols, both genetic- and culture-based, that induce the differentiation of embryonic stem (ES) cells towards a pancreatic beta-cell type. The success of previous protocols in generating insulin-producing beta-cells has been questioned due in part to uncertainty regarding cell lineage but also due to the controversy regarding the source of any insulin detected in these cells. In an attempt to address the latter, we designed a novel assay that can identify de novo insulin synthesis. The method is based on metabolic labeling combined with a modified radio-immunoassay and will routinely detect less than 5 pg/microl of de novo insulin synthesis in lysates from the insulinoma cell line MIN6. This assay failed to detect any newly translated insulin in an ES cell-derived population generated using an adapted version of a previously published, 5-stage differentiation protocol. In combination with other techniques, including immunofluorescent staining and western blot analysis to detect and quantify C-peptide, we conclude that the majority of the insulin found in these differentiated ES cell cultures is medium-derived.
Collections Ireland -> Dublin City University -> PubMed

Full list of authors on original publication

Lorraine O'Driscoll, Martin Clynes, Paul Barham, Finbarr O'Sullivan, Niall W Barron, Eadaoin Mc Kiernan

Experts in our system

1
Lorraine O'Driscoll
Trinity College Dublin
Total Publications: 152
 
2
Martin Clynes
Dublin City University
Total Publications: 209
 
3
Finbarr O'Sullivan
Dublin City University
Total Publications: 21
 
4
Niall Barron
Dublin City University
Total Publications: 44