Introduction and aimFinger-prick related DBS (fpDBS) is a new and innovative specimen collection in clinical practice to develop a screening method for forensic toxicology study [1].Despite this, still very limitedly employed is the fpDBS specimens for capillary electrophoresis (CE) analyses. Transferrin glycoforms are a suitable parameter in alcohol abuse investigation and in particular the carbohydrate deficient transferrin (CDT), defined as the percentage distribution of less glycosilated transferrin, is increased after chronic sustained alcohol intake [2]. The aim of this study was to verify if the use of fpDBS could be suitable for CDT screening CE analysis. MethodHuman serum specimens were collected from volunteer subjects. fpDBS sample collection was performed by pricking the little finger, absorbing the capillary blood drops on DBS cards and letting it to air dry. Once dried, samples required a treatment, which included three easy steps: A) sample resuspension from fpDBS with an acid solution (to remove interfering circulating proteins, mainly haemoglobin); B) removing the paper from the sample solution and pH check (3<4); C) sample neutralization (pH 7-8). The final sample was centrifuged and the supernatant was diluted with an iron rich solution before CE analysis. The CE running buffer was borate (pH 7.9) with diaminobutane (7.5 mmol/L), and the separations were performed within a 30 μm i.d. uncoated fused-silica capillary (length 60cm) using a constant voltage (30kV) and detection at 200 nm. The study was executed analysing fpDBS and parallel serum samples collected from each investigated subject and ÍT levels were measured using both high performance liquid chromatography (HPLC) and CE techniques.Results and conclusionThe comparison between fpDBS and parallel serum CDT level, analysed by the traditional CE and HPLC methods, demonstrated significant correlation r2>0.85. In addition, statistical analysis confirmed that the concentration differences measured in DBS specimens were not relevant. Our results demonstrate how the acid treatment allows analysis in CE despite the small volumes and the large amount of various interfering compounds of whole blood. Therefore, the CE technique use coupled to the fpDBS procedure for CDT analysis expresses a simplified and inexpensive tool designed for use in population screening. In conclusion, the fpDBS CE procedure appear suitable in the forensic alcohol abuse investigations.References[1] CP Stove, AS Ingels, PM De Kesel, WE Lambert. Crit Rev Toxicol. 42 (2012) 230-43.[2] JR Delanghe, ML De Buyzere. Clin Chim Acta. 406 (2009) 1-7.
Finger-prick dried blood spot and capillary electrophoresis: a challenge for alcohol abusers screening analysis
BERTASO, Anna;SORIO, DANIELA;Porpiglia, Nadia Maria;DE PALO, Elio Franco
2015-01-01
Abstract
Introduction and aimFinger-prick related DBS (fpDBS) is a new and innovative specimen collection in clinical practice to develop a screening method for forensic toxicology study [1].Despite this, still very limitedly employed is the fpDBS specimens for capillary electrophoresis (CE) analyses. Transferrin glycoforms are a suitable parameter in alcohol abuse investigation and in particular the carbohydrate deficient transferrin (CDT), defined as the percentage distribution of less glycosilated transferrin, is increased after chronic sustained alcohol intake [2]. The aim of this study was to verify if the use of fpDBS could be suitable for CDT screening CE analysis. MethodHuman serum specimens were collected from volunteer subjects. fpDBS sample collection was performed by pricking the little finger, absorbing the capillary blood drops on DBS cards and letting it to air dry. Once dried, samples required a treatment, which included three easy steps: A) sample resuspension from fpDBS with an acid solution (to remove interfering circulating proteins, mainly haemoglobin); B) removing the paper from the sample solution and pH check (3<4); C) sample neutralization (pH 7-8). The final sample was centrifuged and the supernatant was diluted with an iron rich solution before CE analysis. The CE running buffer was borate (pH 7.9) with diaminobutane (7.5 mmol/L), and the separations were performed within a 30 μm i.d. uncoated fused-silica capillary (length 60cm) using a constant voltage (30kV) and detection at 200 nm. The study was executed analysing fpDBS and parallel serum samples collected from each investigated subject and ÍT levels were measured using both high performance liquid chromatography (HPLC) and CE techniques.Results and conclusionThe comparison between fpDBS and parallel serum CDT level, analysed by the traditional CE and HPLC methods, demonstrated significant correlation r2>0.85. In addition, statistical analysis confirmed that the concentration differences measured in DBS specimens were not relevant. Our results demonstrate how the acid treatment allows analysis in CE despite the small volumes and the large amount of various interfering compounds of whole blood. Therefore, the CE technique use coupled to the fpDBS procedure for CDT analysis expresses a simplified and inexpensive tool designed for use in population screening. In conclusion, the fpDBS CE procedure appear suitable in the forensic alcohol abuse investigations.References[1] CP Stove, AS Ingels, PM De Kesel, WE Lambert. Crit Rev Toxicol. 42 (2012) 230-43.[2] JR Delanghe, ML De Buyzere. Clin Chim Acta. 406 (2009) 1-7.
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