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High sensitivity (zeptomole) detection of BODIPY- labelled heparan sulfate (HS) disaccharides by ion-paired RP-HPLC and LIF detection enables analysis of HS from mosquito midguts.

Maciej-Hulme, Marissa L.; Leprince, Anaëlle C.N.; Lavin, Andre; Guimond, Scott E.; Turnbull, Jeremy E.; Pelletier, Julien; Yates, Edwin A.; Powell, Andrew K.; Skidmore, Mark

High sensitivity (zeptomole) detection of BODIPY- labelled heparan sulfate (HS) disaccharides by ion-paired RP-HPLC and LIF detection enables analysis of HS from mosquito midguts. Thumbnail


Authors

Marissa L. Maciej-Hulme

Anaëlle C.N. Leprince

Andre Lavin

Jeremy E. Turnbull

Julien Pelletier

Edwin A. Yates

Andrew K. Powell



Abstract

The fine structure of heparan sulfate (HS), the glycosaminoglycan polysaccharide component of cell surface and extracellular matrix HS proteoglycans, coordinates the complex cell signalling processes that control homeostasis and drive development in multicellular animals. In addition, HS is involved in the infection of mammals by viruses, bacteria and parasites. The current detection limit for fluorescently labelled HS disaccharides (low femtomole; 10(-15) mol), has effectively hampered investigations of HS composition in small, functionally-relevant populations of cells and tissues that may illuminate the structural requirements for infection and other biochemical processes. Here, an ultra-high sensitivity method is described that utilises a combination of reverse-phase HPLC, with tetraoctylammonium bromide (TOAB) as the ion-pairing reagent and laser-induced fluorescence detection of BODIPY-FL-labelled disaccharides. The method provides an unparalleled increase in the sensitivity of detection by similar to six orders of magnitude, enabling detection in the zeptomolar range (similar to 10(-21) moles; <1000 labelled molecules). This facilitates determination of HS disaccharide compositional analysis from minute samples of selected tissues, as demonstrated by analysis of HS isolated from the midguts of Anopheles gambiae mosquitoes that was achieved without approaching the limit of detection.

Journal Article Type Article
Acceptance Date Feb 1, 2023
Publication Date Feb 2, 2023
Journal Analytical Methods
Print ISSN 1759-9660
Publisher Royal Society of Chemistry
Peer Reviewed Peer Reviewed
Volume 15
Issue 11
Pages 1461-1469
DOI https://doi.org/10.1039/d2ay01803a
Publisher URL https://pubs.rsc.org/en/content/articlelanding/2023/AY/D2AY01803A

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