title: Dual Detection System for Simultaneous Measurement of Intracellular Fluorescent Markers and Cellular Secretion.
abstract: Glucose-stimulated insulin secretion from pancreatic β-cells within islets of Langerhans plays a critical role in maintaining glucose homeostasis. Although this process is essential for maintaining euglycemia, the underlying intracellular mechanisms that control it are still unclear. To allow simultaneous correlation between intracellular signal transduction events and extracellular secretion, an analytical system was developed that integrates fluorescence imaging of intracellular probes with high-speed automated insulin immunoassays. As a demonstration of the system, intracellular [Ca2+] ([Ca2+]i) was measured by imaging Fura-2 fluorescence simultaneously with insulin secretion from islets exposed to elevated glucose levels. Both [Ca2+]iand insulin were oscillatory during application of 10 mM glucose with temporal and quantitative profiles similar to what has been observed elsewhere. In previous work, sinusoidal glucose levels have been used to test the entrainment of islets while monitoring either [Ca2+]ior insulin levels; using this newly developed system, we show unambiguously that oscillations of both [Ca2+]iand insulin release are entrained to oscillatory glucose levels and that the temporal correlation of these are maintained throughout the experiment. It is expected that the developed analytical system can be expanded to investigate a number of other intracellular messengers in islets or other stimulus-secretion pathways in different cells.
authors.affiliation: Department of Chemistry and Biochemistry, Florida State University , 95 Chieftain Way, Dittmer Building, Tallahassee, Florida 32306, United States.
authors.forename: Lian
authors.initials: L
authors.lastname: Yi
authors.affiliation: Department of Chemistry and Biochemistry, Florida State University , 95 Chieftain Way, Dittmer Building, Tallahassee, Florida 32306, United States.
authors.forename: Basel
authors.initials: B
authors.lastname: Bandak
authors.affiliation: Department of Chemistry and Biochemistry, Florida State University , 95 Chieftain Way, Dittmer Building, Tallahassee, Florida 32306, United States.
authors.forename: Xue
authors.initials: X
authors.lastname: Wang
authors.affiliation: Department of Mathematics and Program in Neuroscience, Florida State University , Tallahassee, Florida 32306, United States.|Program in Molecular Biophysics, Florida State University , Tallahassee, Florida 32306, United States.
authors.forename: Richard
authors.initials: R
authors.lastname: Bertram
authors.affiliation: Department of Chemistry and Biochemistry, Florida State University , 95 Chieftain Way, Dittmer Building, Tallahassee, Florida 32306, United States.|Program in Molecular Biophysics, Florida State University , Tallahassee, Florida 32306, United States.
authors.forename: Michael G
authors.initials: MG
authors.lastname: Roper
mesh_terms: D000818:Animals; D002118:Calcium / Q000032:analysis* / Q000378:metabolism; D004867:Equipment Design; D005456:Fluorescent Dyes / Q000032:analysis; D016257:Fura-2 / Q000032:analysis; D005947:Glucose / Q000378:metabolism; D006801:Humans; D007328:Insulin / Q000032:analysis* / Q000378:metabolism; D050417:Insulin-Secreting Cells / Q000378:metabolism; D007515:Islets of Langerhans / Q000378:metabolism; D046210:Microfluidic Analytical Techniques / Q000295:instrumentation*; D061848:Optical Imaging / Q000295:instrumentation*
chemical_list: D005456:Fluorescent Dyes; D007328:Insulin; D005947:Glucose; D002118:Calcium; D016257:Fura-2