Immunoassays have made it possible to measure dozens of individual proteins and other analytes in human samples for help in establishing the diagnosis and prognosis of disease. In too many cases the ...results of those measurements are misleading and can lead to unnecessary treatment or missed opportunities for therapeutic interventions. These cases stem from problems inherent to immunoassays performed with human samples, which include a lack of concordance across platforms, autoantibodies, anti-reagent antibodies, and the high-dose hook effect. Tandem mass spectrometry may represent a detection method capable of alleviating many of the flaws inherent to immunoassays. We review our understanding of the problems associated with immunoassays on human specimens and describe methodologies using tandem mass spectrometry that could solve some of those problems. We also provide a critical discussion of the potential pitfalls of novel mass spectrometric approaches in the clinical laboratory.
Vitamin D deficiency is associated with poor bone health and other adverse health outcomes; however, the associations are greatly attenuated in black vs white individuals. One possible explanation ...for this attenuation is different concentrations of bioavailable vitamin D metabolites in plasma, which are estimated with equations that include the total concentration of vitamin D binding globulin (VDBG) and haplotype-specific dissociation constants.
We developed a method to quantify VDBG with LC-MS/MS that could also identify the haplotypes/isoforms of VDBG present. We validated the method according to recent recommendations for publications of biomarker studies. We determined serum VDBG concentrations in samples from the Atherosclerosis Risk in Communities cohort and compared the results with a widely used monoclonal immunoassay.
With 10 μL of serum or plasma, the lower limit of quantification for the assay (<20% CV) was 71 μg/mL. The assay was linear from 62 to 434 μg/mL, with total imprecision of 7.3-9.0% CV at approximately 250 μg/mL. Significant hemolysis interfered with quantification. The identification of isoforms was 97% concordant with genotyping (κ coefficient). Method comparison with immunoassay revealed significant isoform-specific effects in the immunoassay. Mean concentrations (SD) of VDBG by mass spectrometry were similar in whites and blacks 262 (25) vs 266 (35) μg/mL, respectively; P = 0.43.
Validated mass spectrometric methods for the quantification of proteins in human samples can provide additional information beyond immunoassay. Counter to prior observations by immunoassay, VDBG concentrations did not vary by race.
Cancer genomics research aims to advance personalized oncology by finding and targeting specific genetic alterations associated with cancers. In genome-driven oncology, treatments are selected for ...individual patients on the basis of the findings of tumour genome sequencing. This personalized approach has prolonged the survival of subsets of patients with cancer. However, many patients do not respond to the predicted therapies based on the genomic profiles of their tumours. Furthermore, studies pairing genomic and proteomic analyses of samples from the same tumours have shown that the proteome contains novel information that cannot be discerned through genomic analysis alone. This observation has led to the concept of proteogenomics, in which both types of data are leveraged for a more complete view of tumour biology that might enable patients to be more successfully matched to effective treatments than they would using genomics alone. In this Perspective, we discuss the added value of proteogenomics over the current genome-driven approach to the clinical characterization of cancers and summarize current efforts to incorporate targeted proteomic measurements based on selected/multiple reaction monitoring (SRM/MRM) mass spectrometry into the clinical laboratory to facilitate clinical proteogenomics.
Blood-based indicators that are used in the assessment of iron status are assumed to be accurate. In practice, inaccuracies in these measurements exist and stem from bias and variability. For ...example, the analytic variability of serum ferritin measurements across laboratories is very high (>15%), which increases the rate of misclassification in clinical and epidemiologic studies. The procedures that are used in laboratory medicine to minimize bias and variability could be used effectively in clinical research studies, particularly in the evaluation of iron deficiency and its associated anemia in pregnancy and early childhood and in characterizing states of iron repletion and excess. The harmonization and standardization of traditional and novel bioindicators of iron status will allow results from clinical studies to be more meaningfully translated into clinical practice by providing a firm foundation for clinical laboratories to set appropriate cutoffs. In addition, proficiency testing monitors the performance of the methods over time. It is important that measures of iron status be evaluated, validated, and performed in a manner that is consistent with standard procedures in laboratory medicine.
One of the hallmarks of diabetes is impaired endothelial function. Previous studies showed that HDL can exert protective effects on endothelium stimulating NO production and protecting from ...inflammation and suggested that HDL in obese people with diabetes and dyslipidemia may have lower endothelial protective function. We aimed to investigate whether type 2 diabetes impairs HDL endothelium protective functions in people with otherwise normal lipid profile.
In a case-control study (n = 41 per group) nested in the Cooper Center Longitudinal Study we tested the ability of HDL to protect endothelium by stimulating endothelial nitric oxide synthase activity and suppressing NFκB-mediated inflammatory response in endothelial cells. In parallel we measured HDL protein composition, sphinogosine-1-phosphate and P-selectin.
Despite similar levels of plasma HDL-C the HDL in individuals with type 2 diabetes lost almost 40% of its ability to stimulate eNOS activity (P<0.001) and 20% of its ability to suppress TNFα-dependent NFκB-mediated inflammatory response in endothelial cells (P<0.001) compared to non-T2D controls despite similar BMI and lipid profile (HDL-C, LDL-C, TC, TG). Significantly, the ability of HDL to stimulate eNOS activity was negatively associated with plasma levels of P-selectin, an established marker of endothelial dysfunction (r = -0.32, P<0.001). Furthermore, sphingosine-1-phosphate (S1P) levels were decreased in diabetic plasma (P = 0.017) and correlated with HDL-mediated eNOS activation.
Collectively, our data suggest that HDL in individuals with type 2 diabetes loses its ability to maintain proper endothelial function independent of HDL-C, perhaps due to loss of S1P, and may contribute to development of diabetic complications.
Celotno besedilo
Dostopno za:
DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
Background Efforts to minimize COVID-19 exposure during the current SARS-CoV-2 pandemic have led to limitations in access to medical care and testing. The Tasso-SST kit includes all of the components ...necessary for remote, capillary blood self-collection. In this study, we sought to investigate the accuracy and reliability of the Tasso-SST device as a self-collection device for measurement of SARS-CoV-2 IgG antibodies. Methods Capillary blood was obtained via unsupervised and supervised application of the Tasso-SST device, and venous blood was collected by standard venipuncture. Unsupervised self-collected blood samples underwent either extreme summer or winter-simulated shipping conditions prior to testing. Sera obtained by all three methods were tested concurrently using the EuroImmun anti-SARS-CoV-2 S1 IgG assay in a CLIA-certified clinical laboratory. Results Successful Tasso-SST capillary blood collection by unsupervised and supervised administration was completed by 93.4% and 94.5% of participants, respectively. Sera from 56 participants, 55 with documented (PCR+) COVID-19, and 33 healthy controls were then tested for anti-SARS-CoV-2 IgG antibodies. Compared to venous blood results, Tasso-SST-collected (unstressed) and the summer- and winter-stressed blood samples demonstrated Deming regression slopes of 1.00 (95% CI: 0.99-1.02), 1.00 (95% CI: 0.98-1.01), and 0.99 (95% CI: 0.97-1.01), respectively, with an overall accuracy of 98.9%. Conclusions Capillary blood self-collection using the Tasso-SST device had a high success rate. Moreover, excellent concordance was found for anti-SARS-CoV-2 IgG results between Tasso-SST capillary and standard venous blood-derived sera. The Tasso-SST device should enable widespread collection of capillary blood for testing without medical supervision, facilitating epidemiologic studies.
Celotno besedilo
Dostopno za:
DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
Longitudinal studies testing the relationship between repeated measures of vitamin D or fibroblast growth factor 23 (FGF23) and infectious and cardiac hospitalizations and death in hemodialysis ...patients have not been reported. We examined the association between yearly 25-hydroxyvitamin D (25(OH)D), 1,25-dihydroxyvitamin D (1,25(OH)2D), and FGF23 serum levels and various clinical outcomes using time-dependent Cox regression models with repeated yearly measures and fixed-covariate Cox models with only baseline values after controlling for important clinical covariates in the HEMO study. During a median follow-up of 3 years, 582 of the 1340 participants died, and 499 and 514 participants had a hospitalization or death attributed to infectious and cardiac causes, respectively. Patients in the highest 25(OH)D quartile had the lowest risk of infectious events (hazard ratio HR 0.66 versus the lowest quartile; 95% confidence interval 95% CI, 0.49-0.89), cardiac events (HR, 0.71; 95% CI, 0.53-0.96), and all-cause mortality (HR, 0.46; 95% CI, 0.34-0.62) in time-dependent analyses. No significant associations of 1,25(OH)2D with clinical outcomes were observed in time-dependent or fixed-covariate Cox models. In contrast, the highest FGF23 quartile was associated with a higher risk of infectious events (HR, 1.57 versus the lowest quartile; 95% CI, 1.13-2.18), cardiac events (HR, 1.49; 95% CI, 1.06-2.08), and all-cause mortality (HR, 1.50; 95% CI, 1.07-2.12) in fixed-covariate Cox models. The addition of inflammation markers into the statistical models did not attenuate these associations. Thus, disordered mineral metabolism may affect outcomes in chronic hemodialysis patients.
Effects of Phosphate Binders in Moderate CKD BLOCK, Geoffrey A; WHEELER, David C; KOOIENGA, Laura ...
Journal of the American Society of Nephrology,
08/2012, Letnik:
23, Številka:
8
Journal Article
Recenzirano
Odprti dostop
Some propose using phosphate binders in the CKD population given the association between higher levels of phosphorus and mortality, but their safety and efficacy in this population are not well ...understood. Here, we aimed to determine the effects of phosphate binders on parameters of mineral metabolism and vascular calcification among patients with moderate to advanced CKD. We randomly assigned 148 patients with estimated GFR=20-45 ml/min per 1.73 m(2) to calcium acetate, lanthanum carbonate, sevelamer carbonate, or placebo. The primary endpoint was change in mean serum phosphorus from baseline to the average of months 3, 6, and 9. Serum phosphorus decreased from a baseline mean of 4.2 mg/dl in both active and placebo arms to 3.9 mg/dl with active therapy and 4.1 mg/dl with placebo (P=0.03). Phosphate binders, but not placebo, decreased mean 24-hour urine phosphorus by 22%. Median serum intact parathyroid hormone remained stable with active therapy and increased with placebo (P=0.002). Active therapy did not significantly affect plasma C-terminal fibroblast growth factor 23 levels. Active therapy did, however, significantly increase calcification of the coronary arteries and abdominal aorta (coronary: median increases of 18.1% versus 0.6%, P=0.05; abdominal aorta: median increases of 15.4% versus 3.4%, P=0.03). In conclusion, phosphate binders significantly lower serum and urinary phosphorus and attenuate progression of secondary hyperparathyroidism among patients with CKD who have normal or near-normal levels of serum phosphorus; however, they also promote the progression of vascular calcification. The safety and efficacy of phosphate binders in CKD remain uncertain.
...LC-MS/MS-based clinical protein analysis has predominantly focused on improved analytical measurement for well-established biomarkers (5). Postextraction stability should be determined for both ...pools after storage in the autosampler (^24 h, reinjecting aliquots if feasible), freezing (^72 h, if rou- tine), and extract freeze-thaw for 1 and 2 cycles. Because many preclinical studies rely on biobanked materials, it should be noted that at least 3 freshly acquired samples should be evaluated for stability of 1 freeze-thaw cycle (assay fresh, freeze for ^12 h, thaw for ^2 h, reassay, and compare).
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