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ABSTRACT: Objective
Glucose-stimulated islet insulin or C-peptide secretion experiments are a fundamental tool for studying and assessing islet function. The goal of this work was to develop Ab-based fluorescent homogenous sensors that would allow rapid, inexpensive, near-instantaneous determinations of insulin and C-peptide levels in biological samples.Research design and methods
Our approach was to use molecular pincer design (Heyduk et al., Anal Chem 2008;80:5152-5159) in which a pair of antibodies recognizing nonoverlapping epitopes of the target are modified with short fluorochrome-labeled complementary oligonucleotides and are used to generate a fluorescence energy transfer (FRET) signal in the presence of insulin or C-peptide.Results
The sensors were capable of detecting insulin and C-peptide with high specificity and with picomolar concentration detection limits in times as short as 20 min. Insulin and C-peptide levels determined with the FRET sensors showed outstanding correlation with determinations performed under the same conditions with enzyme-linked immunosorbent assay. Most importantly, the sensors were capable of rapid and accurate determinations of insulin and C-peptide secreted from human or rodent islets, verifying their applicability for rapid assessment of islet function.Conclusions
The homogeneous, rapid, and uncomplicated nature of insulin and C-peptide FRET sensors allows rapid assessment of ?-cell function and could enable point-of-care determinations of insulin and C-peptide.
SUBMITTER: Heyduk E
PROVIDER: S-EPMC3279555 | biostudies-literature | 2010 Oct
REPOSITORIES: biostudies-literature
Heyduk Ewa E Moxley Michael M MM Salvatori Alison A Corbett John A JA Heyduk Tomasz T
Diabetes 20100709 10
<h4>Objective</h4>Glucose-stimulated islet insulin or C-peptide secretion experiments are a fundamental tool for studying and assessing islet function. The goal of this work was to develop Ab-based fluorescent homogenous sensors that would allow rapid, inexpensive, near-instantaneous determinations of insulin and C-peptide levels in biological samples.<h4>Research design and methods</h4>Our approach was to use molecular pincer design (Heyduk et al., Anal Chem 2008;80:5152-5159) in which a pair o ...[more]