Correlative FLIM/Raman using the Renishaw inVia^TM Raman microscope

Session

Poster Session Two

Authors

Dr Dale Boorman (1)

Affiliations

1. Renishaw

Keywords

Raman spectroscopy, fluorescence lifetime imaging (FLIM), multimodal imaging, label-free imaging, correlative microscopy

Abstract text

Fluorescence lifetime imaging (FLIM) is highly compatible with Raman spectroscopy as it utilises autofluorescence, so it can be used to analyse unlabelled samples. Complementary information can be obtained from the two techniques, making correlative FLIM/Raman of significant interest.

In FLIM, an excitation pulse stimulates sample fluorescence, and the time taken for the fluorescence to reach the detector (fluorescence lifetime) is measured. This means that the technique is fast, and it can therefore provide an effective precursor to Raman spectroscopy.

We have integrated FLIM hardware, developed by Becker and Hickl, into our inVia confocal Raman microscope. This enables both Raman and FLIM to be performed on the same system. Both techniques employ the same optical arrangement and mapping stage hardware, meaning that FLIM and Raman data can be collected from the same sample area with pixel-to-pixel accuracy.

We have applied correlative FLIM/Raman within several different application areas. FLIM provides an effective method for initial exploration of samples, with the ability to quickly highlight regions of interest. Raman spectroscopy can then be performed on target areas to understand the underlying chemistry.

Overall, we demonstrate a solution for performing both Raman and FLIM on a single system with pixel-to-pixel correlation. This enables users to collect detailed information about sample structure and composition with ease.