BackgroundVitamin B1 (thiamine-diphosphate) and B6 (pyridoxal-5’phosphate) are micronutrients. Analysis of these micronutrients is important to diagnose potential deficiency which often occurs in elderly people due to malnutrition, in severe alcoholism and in gastrointestinal compromise due to bypass surgery or disease. Existing High Performance Liquid Chromatography (HPLC) based methods include the need for derivatization and long analysis time. We developed an Ultra High Performance Liquid Chromatography Tandem Mass spectrometry (UHPLC-MS/MS) assay with internal standards for simultaneous measurement of underivatized thiamine-diphosphate and pyridoxal-5’phosphate without use of ion pairing reagent.MethodsWhole blood, deproteinized with perchloric acid, containing deuterium labelled internal standards thiamine-diphosphate(thiazole-methyl-D3) and pyridoxal-5’phosphate(methyl-D3), was analyzed by UHPLC-MS/MS. The method was validated for imprecision, linearity, recovery and limit of quantification. Alternate (quantitative) method comparisons of the new versus currently used routine HPLC methods were established with Deming regression.ResultsThiamine-diphosphate and pyridoxal-5’phosphate were measured within 2.5 minutes instrumental run time. Limits of detection were 2.8 nmol/L and 7.8 nmol/L for thiamine-diphosphate and pyridoxal-5’phosphate respectively. Limit of quantification was 9.4 nmol/L for thiamine-diphosphate and 25.9 nmol/L for pyridoxal-5’phosphate. The total imprecision ranged from 3.5–7.7% for thiamine-diphosphate (44–157 nmol/L) and 6.0–10.4% for pyridoxal-5’phosphate (30–130 nmol/L). Extraction recoveries were 101–102% ± 2.5% (thiamine-diphosphate) and 98–100% ± 5% (pyridoxal-5’phosphate). Deming regression yielded slopes of 0.926 and 0.990 in patient samples (n = 282) and national proficiency testing samples (n = 12) respectively, intercepts of +3.5 and +3 for thiamine-diphosphate (n = 282 and n = 12) and slopes of 1.04 and 0.84, intercepts of -2.9 and +20 for pyridoxal-5’phosphate (n = 376 and n = 12).ConclusionThe described UHPLC-MS/MS method allows simultaneous determination of underivatized thiamine-diphosphate and pyridoxal-5’phosphate in whole blood without intensive sample preparation.
In general, the different assays tested for BNP and NT-pro-BNP correlate very well in patients with suspected CHF and may aid in the risk stratification process in emergency departments. However, the value must always be interpreted in conjunction with other clinical information. It should also be considered that renal impairment can affect the results.
Background: During normal pregnancy, the glomerular filtration rate (GFR) increases dramatically. Failure to obtain this physiological increase is an important risk factor for morbidity and mortality for both mother and child. The estimated GFR (eGFR) using serum creatinine levels is unsuitable for accurate measurement of renal function during pregnancy. Therefore, new biomarkers have been proposed. Elevated levels of Cystatin C (CysC) and Neutrophil Gelatinase-Associated Lipocalin (NGAL) are associated with renal failure and preeclampsia (PE). In this study, we determined reference intervals for CysC and NGAL during pregnancy.
Methods: Healthy pregnant women were recruited and blood samples were collected at 9-13 weeks (T1), 27-29 weeks (T2) and 36-39 weeks (T3) of gestation and at 4-13 weeks post-partum (PP). The samples from women with uncomplicated pregnancy were analyzed to determine median values and upper reference limits (URLs, 97.5th percentiles) of creatinine, CysC and NGAL.
Results: A total of 175 women were included. Longitudinal changes and median values of creatinine, CysC and NGAL were determined using only complete data sets (n=59). URLs were determined using all available data. The URL at T1, T2, T3 and PP were 60, 63, 74, 93 µmol/L for creatinine; 0.93, 1.04, 1.61, 1.23 mg/L for CysC; and 87, 84, 88, 95 ng/mL for NGAL.
Conclusions: CysC concentrations are highly dynamic and increase during pregnancy. NGAL concentrations are less dynamic, but well below the URL specified by the manufacturer for non-pregnant women. It is therefore recommended to use trimester-specific reference values for both CysC and NGAL.
The aCCP assay on the IMMULITE 2000(XPi) has good performance characteristics and shows high level of concordance with the aCCP test on Immunoscan RA ELISA test, Immunocap 250 and the Modular systems.
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.