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Quantification by nano liquid chromatography parallel reaction monitoring mass spectrometry of human apolipoprotein A-I, apolipoprotein B, and hemoglobin A1c in dried blood spots.

ABSTRACT: Proteomic analysis of blood proteins in dried blood spots (DBS) is gaining attention as a possible replacement for measurements in plasma/serum collected by venipuncture. We aimed to develop and provisionally validate a nanoflow LC-PRM-MS method for clinical use.We used Skyline to develop a nanoflow LC-PRM-MS method to quantify glycated hemoglobin-?, apolipoprotein A-I, and apolipoprotein B in DBS. Precision, linearity, interferences, and stability were determined and the method was used to analyze samples from 36 human volunteers. The method was compared with clinically validated measurements in paired blood collected via venipuncture.The method was relatively precise for these proteins (10-11% CV) and linear across the normal concentration ranges of these proteins. Interference from high total serum protein concentration (>8 g/dL) was noted for apolipoprotein A-I and apolipoprotein B. Proteins in DBS were stable for 14 days at temperatures below 25°C and trypsinized samples were stable for 48 h at 7°C. There was moderate correlation with clinical methods (r = 0.783-0.858) and significant bias in individual samples.Although the method had adequate precision and linearity for a biomarker, the accuracy compared with clinically validated assays raises concerns regarding the use of DBS compared with venipuncture for clinical use.

SUBMITTER: Henderson CM

PROVIDER: S-EPMC5501987 | biostudies-literature | 2017 Jul

REPOSITORIES: biostudies-literature

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Quantification by nano liquid chromatography parallel reaction monitoring mass spectrometry of human apolipoprotein A-I, apolipoprotein B, and hemoglobin A1c in dried blood spots.

Henderson Clark M CM Bollinger James G JG Becker Jessica O JO Wallace Jennifer M JM Laha Thomas J TJ MacCoss Michael J MJ Hoofnagle Andrew N AN

Proteomics. Clinical applications 20170306 7-8

<h4>Purpose</h4>Proteomic analysis of blood proteins in dried blood spots (DBS) is gaining attention as a possible replacement for measurements in plasma/serum collected by venipuncture. We aimed to develop and provisionally validate a nanoflow LC-PRM-MS method for clinical use.<h4>Experimental design</h4>We used Skyline to develop a nanoflow LC-PRM-MS method to quantify glycated hemoglobin-β, apolipoprotein A-I, and apolipoprotein B in DBS. Precision, linearity, interferences, and stability wer ...[more]

PMID: 28112871

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Project description:BackgroundDolutegravir is currently the preferred component of first-line antiretroviral therapy. To facilitate clinical pharmacology studies in key populations, quantitative analytical methods compatible with microsampling and adaptable to resource-limited settings are desirable. The authors developed and validated a liquid chromatography-ultraviolet detection method to quantify dolutegravir in dried blood spots (DBS).MethodsCalibration standards and quality control samples were prepared by spotting 50 μL of dolutegravir-spiked whole blood on each circle of DBS cards. Three spots (two 6-mm punches/spot) were extracted with methanol. Chromatographic separation was achieved with gradient elution of acetonitrile/potassium phosphate monobasic buffer (pH 5) on a reverse-phase C18 column (flow rate, 1 mL/min) using pioglitazone as the internal standard. UV detection was performed at 260 nm. In the clinical pharmacokinetic study, DBS from finger prick was collected from participants (n = 10) at 8 time points over 12 hours postdosing, with paired plasma at 1 and 12 hours. The method was used to quantify dolutegravir, estimating pharmacokinetic parameters. Agreement between DBS and plasma concentrations was evaluated using linearity and Bland-Altman plots.ResultsThe method was validated over the concentration range of 0.4-10 mcg/mL, accuracy was 102.4%-114.8%, and precision was 3.4%-14.7%. The mean recovery was 42.3% (%CV: 8.3). The mean (±SD) dolutegravir concentration in DBS was 37.5% (±3.8%) lower than that in the plasma. DBS-derived and measured plasma concentrations showed strong correlation with linearity (R2 = 0.9804) and Bland-Altman plots. Means (%CV) of area under curve, Cmax, and C24 from the DBS-derived plasma concentration were 37.8 (23.2) mcg·h/mL, 2.7 (24.7) mcg/mL, and 1.34 (31.6) mcg/mL, respectively.ConclusionsThe application of this simple, accurate, and precise method will expand opportunities for clinical assessment of dolutegravir in resource-limited settings.

Dataset Information

Quantification by nano liquid chromatography parallel reaction monitoring mass spectrometry of human apolipoprotein A-I, apolipoprotein B, and hemoglobin A1c in dried blood spots.

Publications

Quantification by nano liquid chromatography parallel reaction monitoring mass spectrometry of human apolipoprotein A-I, apolipoprotein B, and hemoglobin A1c in dried blood spots.

Similar Datasets

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