{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,9,29]],"date-time":"2026-09-29T16:11:15Z","timestamp":1790698275048,"version":"4.1.0"},"reference-count":40,"publisher":"MDPI AG","issue":"23","license":[{"start":{"date-parts":[[2021,12,3]],"date-time":"2021-12-03T00:00:00Z","timestamp":1638489600000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["Molecules"],"abstract":"<jats:p>Fabry disease (FD) is a rare X-linked lysosomal storage disorder caused by \u03b1-galactosidase A gene (GLA) mutations, resulting in loss of activity of the lysosomal hydrolase, \u03b1-galactosidase A (\u03b1-Gal A). As a result, the main glycosphingolipid substrates, globotriaosylceramide (Gb3) and globotriaosylsphingosine (lyso-Gb3), accumulate in plasma, urine, and tissues. Here, we propose a simple, fast, and sensitive method for plasma quantification of lyso-Gb3, the most promising secondary screening target for FD. Assisted protein precipitation with methanol using Phree cartridges was performed as sample pre-treatment and plasma concentrations were measured using UHPLC-MS\/MS operating in MRM positive electrospray ionization. Method validation provided excellent results for the whole calibration range (0.25\u2013100 ng\/mL). Intra-assay and inter-assay accuracy and precision (CV%) were calculated as &lt;10%. The method was successfully applied to 55 plasma samples obtained from 34 patients with FD, 5 individuals carrying non-relevant polymorphisms of the GLA gene, and 16 healthy controls. Plasma lyso-Gb3 concentrations were larger in both male and female FD groups compared to healthy subjects (p &lt; 0.001). Normal levels of plasma lyso-Gb3 were observed for patients carrying non-relevant mutations of the GLA gene compared to the control group (p = 0.141). Dropping the lower limit of quantification (LLOQ) to 0.25 ng\/mL allowed us to set the optimal plasma lyso-Gb3 cut-off value between FD patients and healthy controls at 0.6 ng\/mL, with a sensitivity of 97.1%, specificity of 100%, and accuracy of 0.998 expressed by the area under the ROC curve (C.I. 0.992 to 1.000, p-value &lt; 0.001). Based on the results obtained, this method can be a reliable tool for early phenotypic assignment, assessing diagnoses in patients with borderline GalA activity, and confirming non-relevant mutations of the GLA gene.<\/jats:p>","DOI":"10.3390\/molecules26237358","type":"journal-article","created":{"date-parts":[[2021,12,6]],"date-time":"2021-12-06T03:10:38Z","timestamp":1638760238000},"page":"7358","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":11,"title":["A Rapid and Simple UHPLC-MS\/MS Method for Quantification of Plasma Globotriaosylsphingosine (lyso-Gb3)"],"prefix":"10.3390","volume":"26","author":[{"ORCID":"https:\/\/orcid.org\/0000-0002-0905-2195","authenticated-orcid":false,"given":"Alessandro","family":"Perrone","sequence":"first","affiliation":[{"name":"IRCCS Istituto delle Scienze Neurologiche di Bologna, 40139 Bologna, Italy"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-3311-1404","authenticated-orcid":false,"given":"Susan","family":"Mohamed","sequence":"additional","affiliation":[{"name":"IRCCS Istituto delle Scienze Neurologiche di Bologna, 40139 Bologna, Italy"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Vincenzo","family":"Donadio","sequence":"additional","affiliation":[{"name":"IRCCS Istituto delle Scienze Neurologiche di Bologna, 40139 Bologna, Italy"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Rocco","family":"Liguori","sequence":"additional","affiliation":[{"name":"IRCCS Istituto delle Scienze Neurologiche di Bologna, 40139 Bologna, Italy"},{"name":"Department of Biomedical and Neuromotor Sciences, University of Bologna, 40139 Bologna, Italy"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Manuela","family":"Contin","sequence":"additional","affiliation":[{"name":"IRCCS Istituto delle Scienze Neurologiche di Bologna, 40139 Bologna, Italy"},{"name":"Department of Biomedical and Neuromotor Sciences, University of Bologna, 40139 Bologna, Italy"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"1968","published-online":{"date-parts":[[2021,12,3]]},"reference":[{"key":"ref_1","unstructured":"Scriver, C.R., Sly, W.A., Beaudet, A.L., and Valle, D. 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