Hilderink M. - Label-Free Detection of Insulin and Glucagon within Human Islets of Langerhans Using Raman Spectroscopy

Janneke Hilderink, Cees Otto, Cees Slump, Aufried Lenferink, Marten Engelse, Clemens van
Blitterswijk, Eelco de Koning, Marcel Karperien, Aart van Apeldoorn

Plos one 2013


Intrahepatic transplantation of donor islets of Langerhans is a promising therapy for patients with type 1 diabetes. It is
of critical importance to accurately monitor islet quality before transplantation, which is currently done by standard
histological methods that are performed off-line and require extensive sample preparation. As an alternative, we
propose Raman spectroscopy which is a non-destructive and label-free technique that allows continuous real-time
monitoring of the tissue to study biological changes as they occur. By performing Raman spectroscopic
measurements on purified insulin and glucagon, we showed that the 520 cm-1 band assigned to disulfide bridges in
insulin, and the 1552 cm-1 band assigned to tryptophan in glucagon are mutually exclusive and could therefore be
used as indirect markers for the label-free distinction between both hormones. High-resolution hyperspectral Raman
imaging for these bands showed the distribution of disulfide bridges and tryptophan at sub-micrometer scale, which
correlated with the location of insulin and glucagon as revealed by conventional immunohistochemistry. As a
measure for this correlation, quantitative analysis was performed comparing the Raman images with the fluorescence
images, resulting in Dice coefficients (ranging between 0 and 1) of 0.36 for insulin and 0.19 for glucagon. Although
the use of separate microscope systems with different spatial resolution and the use of indirect Raman markers
cause some image mismatch, our findings indicate that Raman bands for disulfide bridges and tryptophan can be
used as distinctive markers for the label-free detection of insulin and glucagon in human islets of Langerhans.