There is a need for simple, quantitative and prospective assays for islet quality assessment that are predictive of islet transplantation outcome. The current state-of-the-art athymic nude mouse bioassay is costly, technically challenging and retrospective. In this study, we report on the ability of 2 parameters characterizing human islet quality: (1) oxygen consumption rate (OCR), a measure of viable volume; and (2) OCR/DNA, a measure of fractional viability, to predict diabetes reversal in nude mice. Results demonstrate that the probability for diabetes reversal increases as the graft's OCR/DNA and total OCR increase. For a given transplanted OCR dose, diabetes reversal is strongly dependent on OCR/DNA. The OCR and OCR/DNA (the 'OCR test') data exhibit 89% sensitivity and 77% specificity in predicting diabetes reversal in nude mice (n = 86). We conclude that the prospective OCR test can effectively replace the retrospective athymic nude mouse bioassay in assessing human islet quality prior to islet transplantation.

Despite improvements in outcomes for human islet transplantation, characterization of islet preparations remains poorly defined. This study used both light (LM) and electron microscopy (EM) to characterize 33 islet preparations used for clinical transplants. EM allowed accurate identification and quantification of cell types with measured cell number fractions (mean ± SEM) 35.6 ± 2.1% β-cells, 12.6 ± 1.0% non-β-islet cells, (48.3 ± 2.6% total islet cells), 22.7 ± 1.5% duct cells, and 25.3 ± 1.8% acinar cells. Of the islet cells, 73.6 ± 1.7% were β cells. For comparison to the literature, estimates of cell number fraction, cell volume, and extracellular volume were combined to convert number fraction data to volume fractions applicable to cells, islets, and the entire preparation. The mathematical framework for this conversion was developed. By volume, β cells were 86.5 ± 1.1% of the total islet cell volume and 61.2 ± 0.8% of intact islets (including the extracellular volume), which is similar to that of islets in the pancreas. Our estimates gave 1560 ± 20 cells in an islet equivalent (volume of 150-μm diameter sphere), of which 1140 ± 15 were β cells. To test if LM analysis of the same tissue samples could provide reasonable estimates of purity of the islet preparations, volume fraction islet tissue was measured on thin sections available from 27 of the clinical preparations by point counting morphometrics. Islet purity (islet volume fraction) of individual preparations determined by LM and EM analysis correlated linearly with excellent agreement (R2 = 0.95). However, islet purity by conventional dithizone staining was substantially higher with a 20-30% overestimation. Thus, both EM and LM provide accurate methods to determine the cell composition of human islets preparations and can help us understand many of the discrepancies of islet composition in the literature.

Improvements in pancreatic islet transplantation for treatment of diabetes are hindered by the absence of meaningful islet quality assessment methods. Oxygen consumption rate (OCR) has previously been used to assess the quality of organs and primary tissue for transplantation. In this study, we describe and characterize a stirred microchamber for measuring OCR with small quantities of islets. The device has a titanium body with a chamber volume of about 200 µL and is magnetically stirred and water jacketed for temperature control. Oxygen partial pressure (pO 2 ) is measured by fluorescence quenching with a fiber optic probe, and OCR is determined from the linear decrease of pO 2 with time. We demonstrate that measurements can be made rapidly and with high precision. Measurements with βTC3 cells and islets show that OCR is directly proportional to the number of viable cells in mixtures of live and dead cells and correlate linearly with membrane integrity measurements made with cells that have been cultured for 24 h under various stressful conditions.

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