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Variation in human islet viability based on different membrane integrity stains.

M J Barnett¹, D McGhee-Wilson, A M J Shapiro

¹Clinical Islet Program, University of Alberta, Edmonton, Canada.

Cell Transplantation

|November 30, 2004

Summary

Fluorescein diacetate/propidium iodide (FDA/PI) staining for islet viability shows variability. Alternative stains like SYTO-13/ethidium bromide (SYTO/EB) and calcein AM/ethidium homodimer (C/EthD) offer more reliable human islet viability assessment.

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Area of Science:

Islet cell biology
Cell viability assays
Transplantation research

Background:

Membrane integrity fluorescent staining is crucial for evaluating islet viability.
Islet viability data informs product release criteria and culture condition efficacy.
Variations in viability staining results have been observed with different assays.

Purpose of the Study:

To compare three membrane integrity stains for human islet viability assessment.
To identify potential issues with the standard fluorescein diacetate/propidium iodide (FDA/PI) assay.
To evaluate SYTO-13/ethidium bromide (SYTO/EB) and calcein AM/ethidium homodimer (C/EthD) as alternatives.

Main Methods:

Human islets were isolated from cadaveric pancreata.
Islet viability was assessed using FDA/PI, SYTO/EB, and C/EthD staining.

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Percent viability was calculated for each stain using replicate samples of 50 islets.

Main Results:

SYTO/EB and C/EthD staining yielded nearly identical viability results (approx. 57.6-57.9%).
FDA/PI staining consistently showed elevated viability values compared to SYTO/EB.
Significant discrepancies were observed between FDA/PI scoring and visual islet quality.

Conclusions:

The standard FDA/PI stain may not be optimal for assessing human islet viability due to extracellular fluorescence.
SYTO/EB and C/EthD offer potentially more accurate and reliable alternatives for islet viability assessment.
Accurate islet viability assessment is critical for product release and therapeutic efficacy.