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Maciej-Hulme, M, Leprince, A, Lavin, A, Guimond, S, Turnbull, J, Pelletier, J, Yates, E, Powell, AK and Skidmore, MA (2023) 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. Analytical Methods, 15 (11). pp. 1461-1469. ISSN 1759-9660
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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.
Item Type: | Article |
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Additional Information: | Open Access Article. Published on 02 February 2023. This article is licensed under a Creative Commons Attribution 3.0 Unported Licence. |
Subjects: | Q Science > Q Science (General) Q Science > QD Chemistry Q Science > QD Chemistry > QD415 Biochemistry |
Divisions: | Faculty of Natural Sciences > School of Life Sciences |
Depositing User: | Symplectic |
Date Deposited: | 06 Mar 2023 10:07 |
Last Modified: | 21 Apr 2023 08:38 |
URI: | https://eprints.keele.ac.uk/id/eprint/11986 |