Homogeneous insulin and C-Peptide sensors for rapid assessment of insulin and C-peptide secretion by the islets.

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