Aliquots Of Serum Samples Research Articles

The Potential Risk of Reduced Serum Cholinesterase Activity in COVID-19 Patients Suffering From Cytokine Storm.

Background and objective Several blood biochemical parameters are used to biomonitorcoronavirus disease 2019 (COVID-19), caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Reduced serum cholinesterase (ChE) has been suggested to be a predictive indicator of the severity and outcome of COVID-19 infection. This study aimed to examineserum ChE activity in hospitalized and terminally ill COVID-19 patients with cytokine storm and evaluate the enzyme for the in vitro ChE-inhibitory activity of the organophosphate dichlorvos. Methods We determined the serum ChE activity, electrometrically, among hospitalized COVID-19-cytokine storm patientsand their non-cytokine stormcounterparts. Aliquots of serum samples from healthy volunteers, COVID-19-cytokine storm patients, and non-cytokine storm COVID-19 patients were pooled separately. They were incubated in vitro for 10 minutes with dichlorvos at 0.25 or 0.5 μM.Serum samples from the three groups were subjected to ChE inhibition temporally (5-60 minutes) by 0.25 μM dichlorvos to evaluate the kinetics of enzyme inhibition using steady-state kinetics. Results Of the 165 hospitalized patients with COVID-19, 33 (20%) suffered from the cytokine storm. Serum ChE activity offemale COVID-19 patients with cytokine storm was significantly lower than that of the non-cytokine storm counterparts. Risk analysis of reduced serum ChE activity (≥20%) among the 33 COVID-19 patients with cytokine storm compared to 111 non-cytokine storm COVID-19 patients revealed that the formerwere significantly at risk of reduced enzyme activity. In vitro, dichlorvos at 0.25 μM and 0.5 μM significantly inhibited serum ChE activity in all the groups. The COVID-19-cytokine storm group was the least affected. Dichlorvos at 0.25 μM progressively (5-60 minutes) inhibited serum ChE activity. The inhibition kinetic parameters in COVID-19-cytokine storm patients showed a decreasein the half-life of inhibition (14.54%), inhibition rate (51.46%), and total inhibition time (14.55%). Conclusions Reduced serum ChE in COVID-19 patients with cytokine storm could be adopted as a potential additional laboratory examination tool for bedside risk assessment. The in vitro inhibition profile of serum ChE activity by dichlorvos in COVID-19-cytokine storm patients suggests reduced susceptibility of the enzyme to inhibition. The response of COVID-19 patients to ChE-inhibiting medications should be cautiously evaluated with prior in vitro tests.

Reference intervals for glycated albumin during physiological pregnancy of Europid women: Evidences from a prospective observational study

Background and AimsGlycated albumin (GA) may assess glycometabolic control over a short period of time respect to HbA1c, and its use to screen for gestational diabetes in pregnancy has been suggested. To this regard few data on reference intervals (RI) for GA on Europid women have been collected, only from cross-sectional investigations. Aim of this work has been to collect trimester-specific RI for GA in physiological pregnancies, following a longitudinal prospective study. MethodsForty-five healthy pregnant Europid women have been enrolled for whom a GDM screening test was scheduled at 24–28 weeks, in 5 different Italian centers. Only those negative to the OGTT were included. The women had 4 successive visits at 6–10 weeks of gestation, at 16–18 weeks, at 24–28 weeks and at the end of pregnancy. ALT, AST, total bilirubin, C-reactive protein, cholinesterase, creatinine, GGT, glycated albumin, iron, total serum proteins, transferrin were measured in duplicate on aliquots of serum samples by a central laboratory. ResultsThe RI (2.5–97.5 percentiles) for GA were 11.1–14.8 % (I visit), 10.9–15.6 % (II visit), 10.6–14.1 % (III visit) and 10.7–14.3 % (IV visit). The RI of other biomarkers confirmed previously published data. The RI for serum cholinesterase we present are novel, and were 5049–9906 U/L (Iv), 4212–8965 U/L (IIv), 3518–8470 U/L (IIIv) and 3945–8727 U/L (IVv). ConclusionsTrimester-specific RI are important for using GA and serum cholinesterase in pregnancy. However, considering the high inter-individual variability of both markers, the use of longitudinal interpretations of the individual variations of both proteins during pregnancy should be preferred.

Performance evaluation of TRUPCR® HBV Real-time PCR assay for Hepatitis B virus DNA quantification in clinical samples: report from a tertiary care liver centre.

Quantitative Real-time PCR (qPCR) based Hepatitis B virus (HBV) DNA load estimation is crucial for the initiation of treatment and serves as a strong predictor of liver disease progression in HBV infected individuals. HBV DNA quantification has been ever evolving with the addition of new qPCR based kits on a regular basis. The study was carried with an objective to evaluate the performance characteristics of a commercially available qPCR kit (TRUPCR®, 3B Black Bio Biotech, India Ltd.) and compare with CE approved qPCR kit (Artus HBV Real-time PCR, Qiagen, Germany). 121 HBV infected patients were prospectively enrolled from July to December 2016. Aliquots of serum samples were tested in parallel by TRUPCR® and Artus for HBV DNA levels. Genotype D was most predominant genotype in 36.9% (38/121) of patients followed by genotype A in 14.6% (15/121) patients. Median viral load as seen by Artus was log10IU/ml 3.37 (interquartile range log10IU/ml 2.10-10.89) as compared to TRUPCR® where it was log10IU/ml 3.54 (interquartile range log10IU/ml 2.67-11.52). A very good correlation was seen between the two assays (R2 = 0.964) with a concordance rate of 92.6% (112/121). The TRUPCR® qPCR HBV kit is capable of providing reliable and rapid HBV DNA quantitation and together with its much lower costs, presents itself as a good alternative.

A Multi-Center Assessment of Thyroid Function Test Precision in Chemiluminescence Immunoassay (CLIA) Systems

Background: Chemiluminiscence immunoassay (CLIA) is exclusively pragmatic technology for the analysis of biomarker for diagnosis of thyroid disorders. However, performance characteristics of different chemiluminescence immunoassay (CLIA) systems supplied by different manufacturers in diverse set up for thyroid function test (TFT) has not yet been studied well.
 Objective: Our aim is to evaluate laboratory results by assessment of the reproducibility and repeatability of TFTs in three different diagnostic set up to assure the quality of thyroid hormone assay using chemiluminescence immunoassay (CLIA) instruments: Advia Centaur CP (Siemens), Access 2 (Beckman Coulter) and Liaison (Diasorin).
 Materials and Methods: Among the adult male and female individuals visited for thyroid hormone assay, 51 normal individuals were selected for the study. Three aliquots of serum samples were distributed to assess the reproducibility of three different CLIA equipments operated in three diagnostic centers. Additional three aliquots of serum were analyzed weekly for TFT (fFT3, fFT4 and TSH) to check the repeatability of assay in ADVIA Centaur CP set up. Assay precision was determined by reproducibility and repeatability of test results.
 Results: Results of TFTs of serum samples obtained from three different interlaboratory assays using different CLIA systems have achieved good precision showing minimal variance (P>0.05) and acceptable reproducibility. Results are also precise with adequate repeatability showing minimal variance (P >0.05) obtained from the three different intra-laboratory assays in a single CLIA system using ADVIA Centaur CP by same team.
 Conclusion: Our study elucidates the thyroid hormone assay performance of CLIA systems in three centers, which has shown assay precision with good reproducibility and repeatability of thyroid hormone assay. Thus, the analysis of precision as an essential component of quality control is necessary to deliver precise diagnostic services.

Comparative Profiling of N-Glycans Isolated from Serum Samples of Ovarian Cancer Patients and Analyzed by Microchip Electrophoresis

Ovarian cancer is the fifth leading cause of cancer-related mortalities for women in the United States and the most lethal gynecological cancer. Aberrant glycosylation has been linked to several human diseases, including ovarian cancer, and accurate measurement of changes in glycosylation may provide relevant diagnostic and prognostic information. In this work, we used microchip electrophoresis coupled with laser-induced fluorescence detection to determine quantitative differences among the N-glycan profiles of control individuals and late-stage recurrent ovarian cancer patients prior to and after an experimental drug treatment that combined docetaxel and imatinib mesylate. N-Glycans were enzymatically released from 5-μL aliquots of serum samples, labeled with the anionic fluorescent tag, 8-aminopyrene-1,3,6-trisulfonic acid, and analyzed on microfluidic devices. A 22-cm long separation channel, operated at 1250 V/cm, generated analysis times less than 100 s, separation efficiencies up to 8 × 10(5) plates (3.6 × 10(6) plates/m), and migration time reproducibilities better than 0.1% relative standard deviation after peak alignment. Principal component analysis (PCA) and analysis of variance (ANOVA) tests showed significant differences between the control and both pre- and post-treatment cancer samples and subtle differences between the pre- and post-treatment cancer samples. Area-under-the-curve (AUC) values from receiver operating characteristics (ROC) tests were used to evaluate the diagnostic merit of N-glycan peaks, and specific N-glycan peaks used in combination provided AUCs > 0.90 (highly accurate test) when the control and pretreatment cancer samples and control and post-treatment samples were compared.

Development of a simple enzyme immunoassay for blood haptoglobin concentration in cattle and its application in improving food safety

Abstract Objective To verify the role of haptoglobin, a major acute-phase reactant protein in cattle, as a marker to identify health/disease status in cattle and further assess its potential in improving food safety. Sample Population Serum samples from various cattle groups: clinically normal cattle comprising steers (n = 157) and culled dairy cows (n = 92) before death (antemortem [AM]); retained carcasses (n = 57) railed off the line during postmortem (PM) inspection; and apparently AM normal culled dairy cows (n = 57). Procedure -Efficacy of the simplified monoclonal antibody-based enzyme immunoassay was established by comparing results of haptoglobin tests performed independently on aliquots of serum samples by 3 laboratories. Results Haptoglobin concentration was significantly (P ≤ 0.0001) different between the PM retained carcass group (n = 57) and the AM steer (n = 157) and culled dairy cow (n = 92) groups. In addition, haptoglobin concentration in AM steers (n = 157) and culled dairy cows (n = 92) was significantly (P < 0.0012) different, possibly reflecting a higher percentage of underlying pathologic or inflammatory conditions in animals of the latter group. Evaluation in 3 laboratories of sera from a group of culled dairy cows (n = 57), each laboratory performing a different test procedure, indicated that correlation of haptoglobin concentrations was good between the reported test procedure and the unmodified test and the classical hemoglobin-binding assay that measures peroxidase activity. Conclusion Haptoglobin determination is effective in identifying diseased and healthy cattle. It may be a potentially important tool for application at the farm and slaughterhouse as an aid in improving food safety. (Am J Vet Res 1998;59:1101-1107)

Changes in serum immunoreactive and bioactive growth hormone concentrations in boys with advancing puberty and in response to a 20-hour estradiol infusion.

Acceleration of linear growth during puberty is associated with increased GH secretion, although the relationship between growth and GH is complex. As GH exists as a family of isoforms, some of which may not be identified by immunoassay, there may be alterations in isoform secretion during pubertal maturation that result in increased growth. The changes in serum immunoreactive and bioactive GH concentrations across pubertal maturation were determined in 30 boys, aged 6.5-19.3 yr, with idiopathic short stature or constitutional delay of adolescence. Data were grouped as follows: 1) 6 prepubertal boys with bone age 7 yr or less; 2) 5 prepubertal boys with bone age of more than 7 yr, 3) 10 boys in early puberty; 4) 9 boys with mid- to late puberty. Blood was obtained every 20 min from 2000-0800 h. An equal aliquot of each serum sample was pooled for determination of GH by bio- and immunoassays. The mean serum immunoreactive GH concentration increased from 2.1 +/- 0.3, 1.8 +/- 0.3, and 2.9 +/- 0.5 micrograms/L in groups 1, 2, and 3, respectively, to a peak of 4.6 +/- 0.7 micrograms/L in group 4 (P < 0.05 vs. groups 1-3). The mean serum GH bioactivity was 48 +/- 13 micrograms/L in group 1 and declined to 39 +/- 8 and 31 +/- 3 micrograms/L in groups 2 and 3, increasing to a maximum of 64 +/- 15 micrograms/L in group 4 (P < 0.05 vs. group 3). The ratio of bioactive to immunoreactive GH suggests that the biopotencies of secreted isoforms do not increase during pubertal maturation. The role of E2 in increasing GH secretion was characterized in 8 additional early pubertal boys. Each boy received a saline infusion from 1000-0800 h, followed 1 week later by an infusion of E2 at 4.6 nmol/m2.h. Blood was obtained every 15 min from 2200-0800 h for GH and LH and every 60 min for E2 and testosterone. An equal aliquot of each overnight serum sample was pooled for insulin-like growth factor I (IGF-I) and GH by immuno- and bioassays. The mean serum LH concentration decreased from 5.0 +/- 0.9 to 2.3 +/- 0.6 IU/L (P < 0.01), and the E2 concentration increased from 22 +/- 4 to 81 +/- 26 pmol/L (P < 0.01) during saline and E2 infusions, respectively. Mean serum GH concentrations as measured by immunoassay were similar during both infusions (6.6 +/- 1.4 vs. 9.7 +/- 2.1 micrograms/L; saline vs. E2 infusion, respectively). In contrast, the mean serum GH concentration, as measured by bioassay, decreased from 48 +/- 10 micrograms/L during saline infusion to 16 +/- 3 micrograms/L during E2 infusion (P < 0.05). The mean serum IGF-I concentration also decreased significantly from 116 +/- 17 to 93 +/- 15 micrograms/L (saline vs. E2 infusion, respectively; P < 0.05). Thus, although mean overnight serum GH concentrations increase in late puberty, whether measured by immuno- or bioassay, an acute increase in E2 produces an acute decline in serum GH bioactivity and a lesser decline in the serum IGF-I concentration. These unexpected changes indicate that E2 may affect pubertal growth and GH secretion in a complex or biphasic manner depending on the context in which it is administered.