Pancreatic Islet Cell Biology Core

Objective

The objective of the Islet Cell Biology Core is to provide state of the art support including experimental design, islet isolation, and performance of and/or training in an expansive range of assays for physiological and morphometric assessment of pancreatic islet function and growth. In addition to islet cell biology, we offer support and guidance and enlist unique expertise of interested faculty to help adapt existing technologies to solve unique problems that standard methodologies cannot address utilizing a variety of model systems. We foster collaborative initiatives that allow us to expand the services and support we can offer our users. For example, ties with our Physiology Department promise an expansion of consultation and services to study membrane biophysics critical for understanding normal and diseased islet cells.

The core has the advanced technology and expertise to examine calcium influx by fluorescence imaging (utilizing Fura-2 AM ratiometric calcium dye), metabolic and bioenergetic function with the XFe96 Seahorse Extracellular Flux Analyzer coupled with the BioTek Cytation 5 for Seahorse normalization, and perifusion with or without respirometry for our investigators.

Core Functions

Failure of insulin secreting pancreatic beta cells characterizes the progression of all forms of diabetes. The ICBC is thus positioned to contribute in a significant manner to the basic and translational research activities of the Institute of Diabetes, Obesity and Metabolism (IDOM) at the Perelman School of Medicine of the University of Pennsylvania. The ICBC has developed exceptional expertise in working with human and rodent pancreatic islet tissue, acquiring instrumentation and establishing procedures that are not readily available to the average laboratory.

Services:

We offer a range of services that generally begin with islet isolation from rodent animal models and may be followed by a period of culture by the core for as long as 7 days. Islet hormone secretion can be assessed in static “batch” incubations or by more informative perifusions and expensive immunoassays, routinely for insulin and upon request for glucagon and cAMP. The core may provide these services in an ongoing manner or it may provide critical training to allow the investigator laboratory to perform the experiments independently over time. The core also has the advanced technology and expertise to perform individual islet and single cell fluorescence imaging (Ca2+), respirometry with islet batches using a Seahorse Extracellular Flux Analyzer, perifusion coupled with respirometry, and “closed” respirometry experiments for our investigators.

The ICBC currently is available for the consultation and provides a variety of services, including:

• Rodent islet isolation and culture:
• Culture of human pancreatic islets
• Islet perifusion to measure hormone release**
• Batch incubation experiments to measure hormone release**

Mitochondrial Function:

• Quantitative oxygen consumption of perifused freshly isolated or cultured islet tissue using optical methods (requires 500-750 islets).
• Training and support to measure respiration and extracellular acidification rate (ECAR) measured by the Seahorse Extracellular Flux Bioanalyzer and the companion BioTek Cytation5 system for Seahorse normalization

Intracellular calcium imaging:

• Free intracellular Ca2+ measurements using state of the art fluorescence imaging (Zeiss Observer.Z1 with Zen software).

Intact islets dispersion, cell sorting, and aggregation

• Single cells or cell aggregates/pseudoislets with human islets
• Single cells with mouse islet

Morphometry:

• Provide access and instruction for quantitative imaging of pancreatic islet mass
• Provide instruction for IEQ measurements for islets

Electrophysiology Resource:

• Provide beta-cell Vm measurement and record the basic beta-cell electrogram in response to glucose.
• Perform individual channel analysis in well-defined applications

Consultation:

• To optimize use of core services
• To optimize murine models of beta cell specific gene ablation
• To develop new strategies and technologies to solve particular problems.

Training and Education:

The ICBC provides training for graduate students, postdoctoral fellows and research staff in:

• Pancreatic islet isolation and perfusion techniques (Wei Qin, Jeffrey Roman).
• Basic principles of respiration and oxidative phosphorylation and measurement of oxygen consumption by different methods, including phosphorescence respirometry and oxygen consumption and ECAR by Seahorse (Drs. Doliba and Rozo).
• All aspects of islet cell physiology and biochemistry (Drs. Doliba and Stoffers).

Visit our iLab page for current service options and pricing:

**Note: Hormone assays are submitted to and performed by the Radioimmunoassay/Biomarkers Core and are an additional charge