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Title: Determination of haemoglobin derivatives in aged dried blood spot to estimate haematocrit.
Austin Authors: Zakaria, Rosita;Allen, Katrina J;Koplin, Jennifer J;Crinis, Nick ;De Rosa, Lidia;Roche, Peter;Greaves, Ronda F
Affiliation: Murdoch Children's Research Institute, Parkville, Victoria, Australia
School of Health and Biomedical Sciences, RMIT University, Bundoora, Victoria, Australia
Victorian Clinical Genetics Services, Murdoch Children's Research Institute, Flemington Rd, Parkville, Victoria 3052, Australia
Department of Allergy and Clinical Immunology, Royal Children's Hospital, Parkville, Victoria, Australia
Department of Paediatrics, University of Melbourne, Parkville, Victoria, Australia
Core Laboratory, Pathology, The Royal Children's Hospital, Parkville, Victoria, Australia
Clinical Biochemistry, Austin Pathology, Austin Health, Heidelberg, Victoria, Australia
Issue Date: 26-Jun-2019 2019-01-10
Publication information: Clinical chemistry and laboratory medicine 2019; 57(7): 1026-1034
Abstract: Introduction Dried blood spot (DBS) sample applications now encompass analytes related to clinical diagnosis, epidemiological studies, therapeutic drug monitoring, pharmacokinetic and toxicokinetic studies. Haematocrit (Hct) and haemoglobin (Hb) at very high or low concentrations may influence the accuracy of measurement quantification of the DBS sample. In this study, we aimed to predict the Hct of the punched DBS through primary spectrophotometric estimation of its haemoglobin-derivative (Hb-drv) content. Methods Formic acid solution was used to elute Hb-drv content of 3.2 mm spotted blood from its dry matrix. Direct spectrometry measurement was utilised to scan the extracted Hb-drv in the visible spectrum range of 520-600 nm. The linear relationship between an individual's Hct percentage and Hb-drv concentration was applied to estimate the Hct level of the blood spot. De-identified whole blood samples were used for the method development and evaluation studies. Results The Hb-drv estimation is valid in samples >2 months old. Method validation experiments DBS demonstrate linearity between 82.5 and 207.5 g/L, average coefficient of variation of 3.6% (intra-assay) and 7.7% (inter-assay), analytical recovery of 84%, and a high positive correlation (r=0.88) between Hb-drv and the original whole blood Hct. The Bland-Altman difference plot demonstrates a mean difference of 2.4% between the calculated DBS Hct and the directly measured Hct from fresh whole bloods. Conclusions We have successfully developed a simple Hb-drv method to estimate Hct in aged DBS samples. This method can be incorporated into DBS analytical work-flow for the in-situ estimation of Hct and subsequent correction of the analyte of interest as required.
DOI: 10.1515/cclm-2018-0753
PubMed URL: 30838831
Type: Journal Article
Subjects: dried blood spot
haematocrit correction
Appears in Collections:Journal articles

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