Quantification of the islet product: presentation of a standardized current good manufacturing practices compliant system with minimal variability.
BACKGROUND
Accurate islet quantification has proven difficult to standardize in a good manufacturing practices (GMP) approved manner.


METHODS
The influence of assessment variables from both manual and computer-assisted digital image analysis (DIA) methods were compared using calibrated, standardized microspheres or islets alone. Additionally, a mixture of microspheres and exocrine tissue was used to evaluate the variability of both the current, internationally recognized, manual method and a novel GMP-friendly purity- and volume-based method (PV) evaluated by DIA in a semiclosed, culture bag system.


RESULTS
Computer-assisted DIA recorded known microsphere size distribution and quantities accurately. By using DIA to evaluate islets, the interindividual manually evaluated percent coefficients of variation (CV%; n=14) were reduced by almost half for both islet equivalents (IEs; 31% vs. 17%, P=0.002) and purity (20% vs. 13%, P=0.033). The microsphere pool mixed with exocrine tissue did not differ from expected IE with either method. However, manual IE resulted in a total CV% of 44.3% and a range spanning 258 k IE, whereas PV resulted in CV% of 10.7% and range of 60 k IE. Purity CV% for each method were similar approximating 10.5% and differed from expected by +7% for the manual method and +3% for PV.


CONCLUSIONS
The variability of standard counting methods for islet samples and clinical quantities of microspheres mixed with exocrine tissue were reduced with DIA. They were reduced even further by use of a semiclosed bag system compared with standard manual counting, thereby facilitating the standardization of islet evaluation according to GMP standards.
Department of Immunology, Genetics and Pathology, Uppsala University, Uppsala, Sweden. andrew.friberg@igp.uu.se
Andrew S
AS
Friberg
Heide
H
Brandhorst
Peter
P
Buchwald
Masafumi
M
Goto
Camillo
C
Ricordi
Daniel
D
Brandhorst
Olle
O
Korsgren
D002138:Calibration; D048429:Cell Size; D006801:Humans; D007091:Image Processing, Computer-Assisted; D007515:Islets of Langerhans / Q000166:cytology*; D016381:Islets of Langerhans Transplantation / Q000592:standards*; D008863:Microspheres; D015203:Reproducibility of Results