The pathogenesis of type 1 and 2 diabetes is complex, with immune abnormalities in type 1 and insulin resistance in type 2 diabetes, however, beta cell deficiencies and loss of beta cell mass are a feature of both diseases. The preservation of functional beta cell mass is a goal for both types of diabetes. Optical projection tomography (OPT) has been developed as a powerful imaging tool to map the 3 dimensional distribution of gene and protein expression in intact mouse organs. We aimed to use OPT to study beta cell mass in mouse models of type 1 and type 2 diabetes, and in pancreases of normal and diabetic human organ donors. We used the leptin receptor mutant Leprdb/dbmouse model of type 2 diabetes to study the effects if apoptosis inhibition on beta cell mass. The pro-apoptotic Bcl-2 homology domain 3 (BH3)-only molecule Bim is required for glucose-induced beta cell apoptosis. Using OPT, we observed significantly increased beta cell volume in Bim-deficient Leprdb/db mice mice, which suggested that Bim plays an important role in the loss of islet mass in vivo in type 2 diabetes. NOD8.3 mice are T cell receptor transgenic mice with CD8+ T cells specific for the beta cell antigen IGRP, and they develop accelerated autoimmune diabetes. Using OPT we stained pancreas tissues from NOD8.3 mice with anti-insulin, anti-glucagon and anti-CD3 mAbs to quantify the 3 dimensional progression of insulitis and beta cell destruction. We have collected human pancreas tissues from organ donors to perform OPT and study beta cell mass, size, number and distribution in different lobes of the pancreas to better define the islet structure in diabetic patients. OPT is a powerful tool to study the effects of therapeutic intervention on beta cell mass in both type 1 and 2 diabetes.