Hyperspectral spatially offset Raman spectroscopy in a microfluidic channel.

Spatially offset Raman spectroscopy (SORS) enables one to distinguish chemical fingerprints of top and subsurface layers. In this paper, we apply SORS to a microfluidic two-layer system consisting of transparent liquid in a microchannel as the surface layer and microfluidic PDMS chip material as the sublayer. By using an imaging spectrograph connected to a microscope, we perform hyperspectral SORS acquisitions. Furthermore, the focus position z is translated. Thus, we combine the two methods of hyperspectral SORS and defocusing micro-SORS, which leads to an integral characterization of the layered system. The collected top and subsurface layers of Raman scattering at the optical axis (zero spatial offset) largely depends on the focus position z. However, the spatially offset Raman scattered intensity from the subsurface layer is constant for a large range of focus positions z. We claim that there is potential for internal referencing and alignment reproducibility. We demonstrate these findings experimentally in a microfluidic scenario where a 16 μm deep channel is filled with an aqueous hemoglobin solution. Our observation enables consistent concentration measurements in small-volume liquid samples.