Determination of haemoglobin A1c levels using high-performance liquid chromatography of bloodstains

Abstract

This study aimed to determine haemoglobin A1c (HbA1c) levels in bloodstains shed on glass and fabric surfaces on specified test dates. Blood samples were taken from 26 patients (13 diabetic and 13 non-diabetic). Initial HbA1c levels were detected by using high-performance liquid chromatography (HPLC), and bloodstains were created on both cotton fabric and glass surfaces. Samples were processed at different ages (0, 7, 14, 28 and 56 days) by diluting distilled water and then measuring HbA1c levels by HPLC again. In all stains, HbA1c levels could be determined by using HPLC, but there was a moderate rise in accordance with the age of the stains. A statistically significant difference was found for bloodstains on clothes compared to those on glass surfaces. Receiver operating curve analysis found a sensitivity of 1.0 and specificity of 0.923 (cut-off 6.55) for glass surfaces on the seventh day; a sensitivity of 1.0, a specificity of 0.846 (cut-off 6.45) for clothes on the seventh day; a sensitivity of 1.0 and a specificity of 0.923 (cut-off 6.85) for clothes on the 56th day; and a sensitivity of 1.0 and a specificity of 0.846 (cut-off 7.55) for glass surfaces on the 56th day. In conclusion, this study found that HbA1c levels could be measured with high reliability from forensic bloodstains by using HPLC. Thus, in cases where DNA data banks cannot identify individuals, it would make sense to turn to those who have a medical history of diabetes among the suspects with the results of high HbA1c levels.

Analytical performance evaluation of hemoglobin A1c on an ARKRAY HA-8160 analyzer with newly-developed mobile phase buffer

“…To date, many methods, such as enzymatic assays [ 2 ], capillary electrophoresis [ 3 , 4 ], boronate affinity chromatography [ 5 ], ion-exchange high-performance liquid chromatography (ion-exchange HPLC) [ 6 ] and immunoassays [ 7 ], have been developed and updated. Ion-exchange HPLC assays based on the charge difference principle are among the most commonly used methods for measuring HbA1c in laboratories [ 8 , 9 ]. This analytical system usually includes an automatic glycohemoglobin analyzer, eluents, hemolysis & wash solution, and a chromatography column [ 10 ].…”

The crux of time: A meta-analysis of ex vivo whole blood degradation

“…Although RNA might provide insight for long-term studies and complement the short-term degradation of hemoglobin, they have yet to be combined to estimate bloodstain TSD. Wu et al, 2009;Cavalcanti and Silva, 2019) while massspectrometry (Seok et al, 2018;Rankin-Turner et al, 2019), chromatography (Inoue et al, 1991(Inoue et al, , 1992Andrasko, 1997;Arany and Ohtani, 2011;Acar et al, 2020) and fluorescence spectroscopy (Guo et al, 2013;Mc Shine et al, 2017;Weber et al, 2021) have measured changes in metabolites and protein composition of deposited bloodstains. DNA (Kohlmeier and Schneider, 2012;Sirker et al, 2013Sirker et al, , 2016Rahikainen et al, 2016;Stotesbury et al, 2020;Cossette et al, 2021), total RNA, mRNA and miRNA (Bauer et al, 2003;Anderson et al, 2005Anderson et al, , 2011Simard et al, 2012;Qi et al, 2013;Alshehhi et al, 2017;Fang et al, 2019;Fu and Allen, 2019;Mayes et al, 2019;Alshehhi and Haddrill, 2020;Asaghiar and Williams, 2020;Salzmann et al, 2021b;Heneghan et al, 2021;Li et al, 2021;Manasatienjig and Nimnual, 2021;Zhao et al, 2021) degradation have also been targeted in longer time series.…”

The crux of time: A meta-analysis of ex vivo whole blood degradation

“…Photography 13,14,44 and hyperspectral imaging [45][46][47] have measured the colour changes in blood while electron spin resonance and electrochemistry analyzed the conformational changes of the heme groups [48][49][50] . Atomic force microscopy has investigated the changes in RBC physiology over time [51][52][53] while mass-spectrometry 54,55 , chromatography [56][57][58][59][60] and fluorescence spectroscopy [61][62][63] have measured changes in metabolites and protein composition of deposited bloodstains. DNA 38,39,[64][65][66][67] , total RNA, mRNA and miRNA [10][11][12][68][69][70][71][72][73][74][75][76][77][78][79][80] degradation have also been targeted in longer time series.…”