Chemstrips

Individual animal medical observations were recorded daily throughout the study. Observations included appetite, bowel movements, body weight, blood glucose (Accu-Chek Simplicity BG monitor and Chemstrips; maximum value of 600 mg/dL: Roche Diagnostics), urine glucose, and insulin therapy.

The urine glucose concentration of mice was measured weekly using Chemstrips (Roche, Boehringer Mannheim). Diabetes was defined as glycosuria above 500 mg/dL on 2 consecutive tests. For histological analysis, pancreata were harvested from 12-week-old female NOD mice and fixed overnight with 10% formalin, according to the procedure previously described (6 (link)). The severity of insulitis was scored blindly on sections stained with H&E.

Urine glucose concentrations (glycosuria) were measured weekly using Chemstrips (Boehringer Mannheim, Indianapolis, IN, USA). Mice with urine glucose concentration >27.75 mmol/L on two consecutive tests were defined as diabetic. Blood glucose concentrations were monitored daily by Roche ACCU-CHEK (Roche Ltd., Basel, Switzerland) after islet transplantation. Graft rejection and loss of function were defined as blood glucose levels higher than 300 mg/dL for two consecutive days. For the assessment of insulitis, pancreatic tissues were obtained from 14-week-old PBS-treated or ALA-treated NOD mice and the severity of insulitis was scored on haematoxylin–eosin stained sections (Sigma-Aldrich) and classified as described [52 (link)]. The degree of insulitis in the pancreas was evaluated by scoring 15–30 islets/mouse in a blinded fashion according the following criteria: intact islet: no mononuclear cell infiltration; peri-insulitis: mononuclear cell infiltration in <25%; intra-insulitis: mononuclear cell infiltration in 25–50% of the islet; severe insulitis: mononuclear cell infiltration in 50–75% of the islet; destructive insulitis: >75% of the islet was infiltrated.

To evaluate spontaneous diabetes frequency, urine glucose of female transgenic and littermate control mice was measured twice a week using Chemstrips (Boehringer Mannheim, Indianapolis, IN, USA). Mice with a urine glucose concentration >500 mg/dl in two consecutive tests were considered diabetic. For insulitis assessment, pancreases were collected from 12- to 15-week-old mice and fixed in 10% buffered formalin. Hematoxylin- and eosin-stained pancreas sections were blindly scored as described previously (17 (link)).