Poster Presentation HUPO 2019 - 18th Human Proteome Organization World Congress

Multiplexed glycan profiling of extracellular vesicles using lectin-surface enhanced raman spectroscopy (lectin-SERS) (#729)

Edward S.X. Moh 1 , Wei Zhang 1 , Mingkai Wu 1 , Yuling Wang 1 , Nicolle H Packer 1
  1. ARC Centre of Nanoscale BioPhotonics, Macquarie University, Sydney, NSW, Australia

Aberrant cellular glycosylation has been a reported feature of multiple disease states with high-resolution separation and mass spectrometric methodologies being key to these discoveries. However, clinical translation of these discoveries is hard to implement due to reliance on instrumentation and expert data interpretation. The only glycan related clinical example to-date uses the lectin, Lens culinaris agglutinin, to detect fucosylated alpha-fetoprotein-L3 as a marker for hepatocellular carcinoma progression. Differential lectin binding to specific glycan structures can simplify the complexity of detailed mass spectrometric profiling and aid in the clinical application of glyco-oriented disease monitoring and diagnosis.

Multi-lectin profiling provides an advantage, as differences between the glycome of different diseased cells can be subtle and require a panel of lectins for differentiation. Multiplexed fluorescence detection of lectins is limited by the overlapping spectrometric properties of fluorophores, and existing lectin arrays cannot be considered true multiplexing but more about testing lectin binding individually in a standardised or high throughput fashion. Surface enhanced Raman spectroscopy (SERS) using reporter coated gold nanoparticles provides an effective way for multiplexing, as a single excitation wavelength can read out multiple unique Raman reporter signals from different SERS particles coated with diverse targeting probes.

In this work, we developed multiple lectin conjugated SERS-particles and demonstrate the potential of multiplexed lectin profiling of CD63+ extracellular vesicles in vitro and in body fluids, with detection using a handheld Raman reader. We show that enzymatic desialylation of the vesicles reduces the WGA-SERS signal that corresponds to sialic acid binding, with a concurrent increase in PHA-L-SERS signal corresponding to an increase in the exposed galactose terminal sugars; validating that differences in vesicle glycosylation can be detected using this method. Further optimisation and testing of this method will lead to rapid profiling of clinical samples as a pre-screening method for more detailed diagnostic tests.