ICP-MS establishes blood lead reference method expected to improve detection accuracy / 中华检验医学杂志

Lead poisoning severely threatens human health with its cumulation and durability in the body. The analysis of lead in blood is vital for screening, diagnosis, treatment, and prognostication of lead poisoning and for indirectly monitoring the level of lead in the environment. Although the detection programs are available throughout our country, the accuracy and comparability of the results cannot meet the expectation. A variety of factors can affect the accuracy of blood lead testing. To promote the application of blood lead analysis in clinical trials and reduce the bias of results, a better reference system for blood lead analysis should be established to evaluate the accuracy of traditional methods, promote the standardization of blood lead analysis and achieve accurate blood lead testing.

Method and application of high efficient preparation of procalcitonin standby reference material based on protein recombination technology / 中华检验医学杂志

Objective:To construct a prokaryotic expression vector for human retinol binding protein 4 (hRBP4) that allows technicians to obtain hRBP4 purified protein with low cost, high efficiency, high concentration and high purity.Methods:The hRBP4 coding sequence provided by National Center for Biotechnology Information was optimized by E. coli codons, and a synthetic DNA fragment was cloned into the PET-28A (+) prokaryotic expression vector to construct a recombinant hRBP4 expression plasmid. The recombinant protein was transformed into E. coli BL21, and the induced expression conditions (temperature, rotate speed and isopropyl β-d-thiogalactoside concentration) were optimized. The recombinant protein was purified by His fusion tag. Results:The recombinant hRBP4 prokaryotic expression plasmid was successfully constructed, and the expression concentration and induction temperature of the recombinant protein were optimized. The results of sodium dodecyl sulfate polyacrylamide gel electrophoresis showed that a band with a relative molecular weight of 26 000 daltons was clearly visible in the purified product. The purified hRBP4 protein could be detected clinically, and there was a good linear relationship between the dilution ratio and the detection concentration.Conclusions:The recombinant hRBP4 protein has high purity, high concentration, and short production cycle. It has the potential to become a candidate for reference materials for laboratory quality evaluations.

Establishment and validation of 18F-FET PET radiomic features-based model in predicting IDH1 genotype in gliomas / 中华核医学与分子影像杂志

Objective:To establish O-(2-[ 18F]fluoroethyl)- L-tyrosine( 18F-FET) PET radiomics features-based model and investigate its predictive efficacy for isocitrate dehydrogenase type 1 (IDH1) genotyping in untreated gliomas. Methods:From November 2017 to February 2019, 58 pathologically confirmed glioma patients (36 males, 22 females; age (41.8±15.1) years) with preoperative 18F-FET PET/CT imaging in Huashan Hospital, Fudan University were retrospectively enrolled. PyRadiomics software package was used to extract 105 radiomics features. Least absolute shrinkage and selection operator (LASSO) algorithm with 5-fold cross-validation was used to build the logistic regression model. And radiomic scores (RS) of each lesion were calculated according to their weighted coefficients. The area under the receiver operating characteristic (ROC) curve was used for evaluating the predictive efficacy for IDH1 prediction. The predictive efficacies of radiomics model and traditional semi-quantitative parameters including tumor-to-background ratio (TBR; maximum TBR (TBR max), mean TBR (TBR mean), peak TBR (TBR peak)), metabolic tumor volume (MTV) and total lesion tracer uptake (TLU), were compared by Delong test. Results:Seven radiomics features including maximum 2-dimensional (2D) diameter slice, first order_maximum, first order_range, gray level co-occurrence matrix (GLCM)_joint energy, GLCM_inverse variance, gray level dependence matrix (GLDM)_dependence entropy and GLDM_large dependence low gray level emphasis were selected for the LASSO regression model building and RS calculation. ROC analysis results showed that the predictive accuracy of RS for IDH1 genotyping (mutation, n=20; wild-type, n=38) was 81.0%(47/58), with sensitivity of 65.0%(13/20), specificity of 89.5%(34/38), and area under curve (AUC) of 0.842, respectively. The traditional 18F-FET semi-quantitative parameter TLU ranked the second regarding the diagnostic performance, with accuracy of 60.3%(35/58), sensitivity of 85.0%(17/20), specificity of 47.4%(18/38), and AUC of 0.661( z=3.426, P<0.01). Conclusion:Radiomics analysis based on 18F-FET PET images can improve the predictive efficacy for IDH1 genotyping in untreated adult glioma patients.

Study on the mechanism of proliferation-inhibiting effect of icaritin on hepatoma cells via regulating miRNA-329 and miRNA-1236 / 肿瘤研究与临床

Objective:To investigate the molecular mechanism of proliferation-inhibiting effect of icaritin on hepatoma cells via regulating miRNA-329 (miR-329) and miRNA-1236 (miR-1236).Methods:Hepatoma cell line HepG2 was treated with icaritin at different concentrations (2.5, 5.0, 10.0, 20.0, 40.0 μmol/L), and the control group only added dimethyl sulfoxide (DMSO). The half inhibitory concentration ( IC50) of icaritin on HepG2 cells and cell proliferation rate were detected by CCK-8 after cultured for 36 h. HepG2 cells were treated with 400 μg/L alpha fetoprotein (AFP). After cultured for 0, 12, 24, 36, 48 and 60 h, the effect of AFP on the proliferation of HepG2 cells was detected by CCK-8 method. AFP-3'UTR reporter plasmid pmirGLO-AFP-3'UTR plasmid was constructed, pmirGLO blank vector plasmid, pmirGLO-AFP-3'UTR plasmid, miR-329 or miR-1236 mimics or inhibitors, control plasmid of mimics (NC), control plasmid of inhibitors (INC) were respectively co-transfected with HepG2 cells, and the effect of miR-329 and miR-1236 on the luciferase activity was detected by dual-luciferase reporter assay after cultured for 24 h. Western blot and real-time fluorescence quantitative polymerase chain reaction (qRT-PCR) were used to detect the effects of icaritin on the expressions of AFP, miR-329 and miR-1236 in HepG2 cells. HepG2 cells were respectively transfected with mimics and inhibitors of miR-329 and miR-1236 to detect the effects of miR-329 and miR-1236 on the expression of AFP. Results:The cell proliferation rates of 2.5, 5.0, 10.0, 20.0, 40.0 μmol/L icaritin group and control group were (80.4±2.3)%, (73.2±1.6)%, (51.7±3.3)%, (38.2±4.6)%, (29.5±4.3)%, and (94.0±2.9)%, respectively, and the difference was statistically significant ( F = 75.65, P < 0.01); the differences in the proliferation rate of HepG2 cells between different concentrations of icaritin group and control group were statistically significant (all P < 0.01). The IC50 of icaritin on HepG2 cells was 10 μmol/L. The relative expressions of AFP mRNA in 2.5, 5.0, 10.0, 20.0, 40.0 μmol/L icaritin group and control group were 0.83±0.06, 0.69±0.02, 0.53±0.07, 0.45±0.01, 0.33±0.07, and 1.00±0.01, respectively, and the difference was statistically significant ( F = 42.67, P < 0.01); the differences in the relative expressions of AFP mRNA between different concentrations of icaritin group and control group were statistically significant (all P < 0.01). HepG2 cells were treated by 400 μg /L AFP for 0, 12, 24, 36, 48 and 60 h, and the cell proliferation rates were (102±5)%, (138±13)%, (186±24)%, (260±12)%, (311±15)%, and (348±25)%, respectively, and the difference was statistically significant ( F = 27.483, P < 0.01); the differences in the cell proliferation rate between different time of AFP treatment and 0 h were statistically significant (all P < 0.01). Compared with the control group, different concentrations of icaritin can promote the expression of miR-329 and miR-1236 (all P < 0.01). After co-transfection of miR-329 and miR-1236 mimics and AFP-3'UTR, the luciferase activity decreased by about 40%; after co-transfection of miR-329 and miR-1236 inhibitors and AFP-3'UTR, the luciferase activity increased about 1.5 times. Both miR-329 and miR-1236 can reduce the expression levels of AFP protein and mRNA (both P < 0.05). Conclusion:Icaritin can promote the binding of miR-219, miR-1236 and AFP-3'UTR by promoting the expression of miR-329 and miR-1236, inhibit the stability and translation activity of AFP mRNA, inhibit the expression of AFP, and thus inhibit the proliferation of hepatoma cells in vitro.

Establishment of a candidate reference method for plasma normetanephrine and its application in the external quality assessment program / 中华检验医学杂志

Objectives:To establish a candidate reference method of isotope dilution liquid chromatography tandem mass spectrometry (ID-LC/MS/MS) for the determination of human plasma normetanephrine, and to evaluate the performance of the method. The method was used to quantify the samples of the external quality assessment program, and to initially evaluate the detection status of plasma normetanephrine.Methods:The isotope standard solution of normetanephrine was selected as the internal standard, the gravimetric method was used for sampling, and the standard curve method was used for quantification. Protein precipitation combined with weak cation solid phase extraction was used for pretreatment, and ultra-high liquid chromatography-coupled triple quadrupole mass spectrometry was used for LC/MS analysis. According to the relevant EP documents, the specificity, matrix effect, detection limit, quantification limit, precision, accuracy, and uncertainty of the method were estimated. This method is used to quantify the samples of the 2020 National Center for Clinical Laboratories external quality assessment program of normetanephrine. Taking the average value of this method as the target value, the optimal allowable total error standard of biological variation as the evaluation limit, the quality of the laboratory testing was evaluated.Results:The method had good specificity, and the interferences and matrix effects did not affect the detection results. The detection limit and quantification limit of plasma normetanephrine were 1.08 pg/g and 3.54 pg/g, respectively. The intra-batch coefficient of variation ( CV) and total CV were 0.43%-1.10% and 0.61%-1.42%, respectively. The relative recovery rates were 98.5%～101.9%. The relative expansion uncertainty of the four plasma samples were 3.10%, 2.34%, 2.16%, and 1.73%, respectively. The results of the external quality assessment program showed that the pass rates of the 202013 and 202014 samples were 80% and 85%, respectively. Conclusions:The study established a candidate reference method of ID-LC/MS/MS for the measurement of plasma normetanephrine. The method is accurate, precise and simple, and is expected to be used as a reference method for the determination of plasma normetanephrine, and can be applied to quantify the samples of the external quality assessment program.

Establishing allowable total error for serum total folate in external quality assessment / 中华检验医学杂志

Objective:To establish the allowable total error (TEa) of the national external quality assessment (EQA) program in line with the current quality level of serum folate measurement in China.Methods:The data of serum total folate test in the clinical laboratory of a hospital in Beijing in 2016 were collected, and the Stata SE 15 software was used for Monte Carlo simulation to obtain the false-negative rate under different bias and inaccuracy conditions. The Origin Pro 9.1 software was used to make the contour figure. The TEa of serum total folate test is derived based on the acceptable false-negative rate. National EQA data of serum total folate in 2020 were collected to calculate the pass rate of participating laboratories and the laboratory pass rate of quality control products at each level under the five TEa derived from the analysis performance on clinical outcomes, biological variation, and the evaluation criterion of national EQA.Results:Based on the influence of analytical performance on clinical outcomes, the TEa was 10%. Under this TEa, the pass rate of the first EQA program of serum total folate in 2020 was more than 80%, and the pass rate of the second time was 73.1%. Under the minimum (46.57%) and appropriate level of TEa (15.52%) derived from biological variation and national EQA evaluation criterion, the pass rate of serum total folate in the two EQA programs in 2020 exceeded 85%.Conclusion:The analytical performance of serum total folate in China cannot meet the requirements of TEa derived based on the effect of analytical performance on clinical outcomes. An appropriate level of TEa derived based on biological variation (15.52%) is suggested as the recommended criterion for the TEa of serum total folate test.

Current status and standardization conception of the testing quality of therapeutic drug monitoring samples / 中华检验医学杂志

Accurate determination of drug concentration in blood samples is a necessary prerequisite for therapeutic drug monitoring (TDM) and the implementation of precise drug treatment, and it is also one of the important tasks of clinical laboratories.TDM plays an important role in clinical treatment in immunosuppressants (cyclosporine A, tacrolimus), psychotropic drugs (valproic acid, carbamazepine) and other drugs that require monitoring of drug concentration. There are many types of methods used for TDM for the detection of drug concentration in blood samples. At present, only a few immuno-assay methods were approved for marketing with detection systems and kits, most methods used for TDM are high performance liquid chromatography or liquid chromatography-tandem mass spectrometry which belong to laboratory developed tests (LDTs). Detection of TDM samples has many problems, such as incomparable testing results and large bias between different testing systems, including the biasbetween both different methods and different laboratories using the same method. There are several reasons:(1) the traceability chain has not been established, (2) the methods have not yet been standardized, (3) the coverage of the EQA plan is insufficient, (4) the awareness of TDM laboratories to participate in the EQA plan is insufficient, (5) TDM standardization is still in its infancy. These problems restrict the clinical application of TDM and the development of related research work. In order to solve these problems, it is necessary to: (1) Establish a reference system to realize the traceability of the test results; (2) While gradually increasing the TDM EQA plan items, the Trueness evaluation plan should be carried out as soon as possible; (3) Standardized TDM sample testing Technology; (4) Strengthen laboratory management and establish a complete quality management system.

Commutability assessment of 16 processed materials for 17-hydroxyprogesterone based on two approaches in China / 中华检验医学杂志

Objective@#The aim of this study is to evaluate the commutability of 16 processed materials for 17-hydroxyprogesterone by using 2 commutability assessment approaches.@*Methods@#52 serum specimens were collected in Clinical Laboratory Department of Beijing Hospital from February 2018 to June 2019. According to the report of the Clinical and Laboratory Standards Institute (EP14-A3) document and the recommendations of the International Federation of Clinical Chemistry and Laboratory Medicine (IFCC) working group on commutabilityassessment, serum 17-hydroxyprogesterone isotope diluent chromatogram tandem mass spectrometry (ID-LC/MS/MS) was used for comparison. Three clinical routine analysis systems (1 radioimmunoassay, 2 LC/MS analysis methods) were used to determine the concentration of 17-hydroxyprogesterone in 52 human serum samples and 16 processed materialsfor commutabilityassessment.@*Results@#Combined with the results of the two commutability assessment, all accuracy verification materials and national steroid hormone standards showed good commutability in the LC/MS analysis system, and 6/9 EQA materials showed commutability in the three routine analysis systems.All materials showed good commutability in the LC/MS analysis system of bias difference method.@*Conclusions@#The two kinds of commutability assessment results are different. Bias difference method has more clinical value, but it has certain application limitations. The use of fresh frozen human serum as a quality assessment materialfor serum 17-hydroxyprogesterone is meets the commutability requirement.

Preoperative 11C-methionine PET imaging in glioma grading efficacy and its predictive value for IDH1 gene mutation status / 中华核医学与分子影像杂志

Objective:To assess the preoperative 11C-methionine ( 11C-MET) PET imaging in glioma grading efficacy and its predictive value for isocitrate dehydrogenase enzyme 1 (IDH1) gene mutation status. Methods:A total of 118 glioma cases (70 males, 48 females; median age 45 years, age range: 10-71 years; Ⅱ grade 65 cases, Ⅲ grade 34 cases, Ⅳ grade 19 cases) received 11C-MET PET imaging in PET Center of Huashan Hospital from February 2012 to November 2017 were retrospectively analyzed. Lesion-based semi-quantitative analysis was conducted on the 11C-MET imaging. Maximum standardized uptake value (SUV max), peak standardized uptake value (SUV peak), tumor-to-background ratio (TBR; SUV max in lesion/mean standardized uptake value (SUV mean) in normal contralateral cortex) were calculated. Independent-sample t test and one-way analysis of variance were applied to assess the differentiating efficacy of 11C-MET PET imaging for different glioma groups. Based on IDH1 immunohistochemical staining results, predictive efficacy of 11C-MET PET diagnostic parameters on IDH1 mutation status in glioma patients was further analyzed with receiver operating characteristic (ROC) curve analysis. Results:Low-grade glioma (LGG; grade Ⅱ) group showed significant differences from high-grade glioma (HGG; grade Ⅲ-Ⅳ) group in SUV max(2.458±1.100 vs 3.828±1.540; t=5.624, P<0.01), SUV peak (2.160±0.991 vs 3.261±1.319; t=5.175, P<0.01) and TBR (2.283±0.942 vs 3.434±1.395; t=5.328, P<0.01). SUV max (2.458±1.100, 3.591±1.611 and 4.251±1.343; F=17.67, P<0.01), SUV peak(2.160±0.991, 3.040±1.335 and 3.656±1.225; F=15.48, P<0.01) and TBR (2.283±0.942, 3.010±1.242 and 4.192±1.358; F=22.73, P<0.01) were different in grade Ⅱ, Ⅲ and Ⅳ glioma subgroups. SUV max, SUV peak and TBR all showed significant differences between grade Ⅱ and grade Ⅲ gliomas, grade Ⅱ and grade Ⅳ gliomas, and there were also statistical differences between grade Ⅲ and grade Ⅳ glioma with TBR (all P<0.01). SUV max indicated the best single-parameter prediction performance (area under curve (AUC) =0.808, z=7.193, P<0.01), while the SUV max + SUV peak showed the best performance (AUC=0.852, z=9.115, P<0.01). In the subgroup of grade Ⅱ ( n=55), TBR of patients with IDH1 gene mutation ( n=41) was lower than that of patients with IDH1 wild-types ( n=14; 2.152±0.759 vs 2.793±1.208; t=2.326, P=0.02), while TBR of those with oligodendrogenic components ( n=26) was higher than that of patients with IDH1 gene mutation only ( n=18; 2.383±0.825 vs 1.854±0.478; t=2.447, P=0.02). Conclusions:Preoperative semi-quantitative parameters (SUV max, SUV peak, TBR) of 11C-MET brain PET imaging have satisfactory grading discrimination performance for glioma patients. SUV max is the best predictor for IDH1 mutation as a single parameter, while SUV max + SUV peak showed the most optimized predictive ability. The oligodendrogenic components in glioma can increase the uptake of 11C-MET, which may affect the effectiveness of 11C-MET in determining glioma grade to some extent.

Establishment of a candidate reference method for serum glycated albumin based on liquid chromatography tandem mass spectrometry / 中华检验医学杂志

Objective:To establish an accurate quantitative method for the determination of serum glycated albumin (GA) based on isotope dilution liquid chromatography tandem mass spectrometry (ID-LC/MS/MS).Methods:This study optimized the ID-LC/MS method recommended by the Japan Society of Clinical Chemistry (JSCC). Albumin (Alb) was purified by anion exchange chromatography. The standard solution of purified sample, lysine (Lys) and deoxyfructosyl lysine (DOF-Lys) and their mixed internal standard were weighed accurately by weight method. The Alb was hydrolyzed to amino acid, and Lys and DOF-Lys were detected to determine GA. The method was evaluated and compared with the routine method.Results:The purity of Alb was 100%. The R 2 ranges of Lys and DOF-Lys linear standard curves were 0.997 8-0.999 9 and 0.999 4-1.000 0. The results of three-day precision evaluation of low, medium and high concentrations of serum were 0.69%-1.97%, 1.13%-2.33% and 1.37%-2.54% in intra, inter and total Coefficient of variation ( CV). In the accuracy evaluation, the deviations between the results of three concentration standards and the certified value were -2.56%, -1.87% and-2.94%. The matrix effects of Lys and DOF-Lys were 89.92% and 109.98%, and the carryover rates were -1.13% and -3.27% respectively. The detection limit and quantitative limit of Lys were 0.075 nmol/g and 0.248 nmol/g, and the DOF-Lys were 0.007 nmol/g and 0.024 nmol/g. The relative expanded uncertainty was 1.60%-2.03%. The fitting regression curve R 2 with the routine method was 0.970 7. Conclusions:A method was established for the detection of serum GA. The method evaluation was accurate and reliable. It was expected to be a candidate reference method for the detection of serum GA.

A candidate reference method using ICP-MS for simultaneous determination of iron and copper in human blood / 中华检验医学杂志

Objective:To establish a candidate reference method for simultaneous determination of whole blood iron and copper based on ICP-MS technology.Methods:Cobalt (Co) was chosen as the internal standard, and was added gravimetrically into the standard solution and the whole blood sample. The whole blood sample was digested by electric heating plate and diluted. The isotopic ratios of 57Fe/ 59Co and 63Cu/ 59Co were measured by ICP-MS in the isotopic analysis mode. The precision, standard recovery and accuracy of the method were verified by measuring EQA materials and certified standard materials, and the performance of the method was further evaluated by method comparison. Results:The detection limit of iron is 0.136 mg/kg and the limit of quantification was 0.454 mg/kg in this method. The detection limit of copper in whole blood was 0.008 mg/kg and the limit of quantification was 0.027 mg/kg. The standard curve of copper ranges from 0.83-3.33 mg/kg, iron ranges from 167-667 mg/kg. The calibration curve of iron and copper was good linearity ( R 2>0.999 90). The precision of the method did well. The total CV range was 0.42%-0.68% for iron and 0.14%-0.94% for copper. The spike recoveries were all around 100%, the range of iron: 99.69%-100.07%; copper: 99.26%-100.51%. The correlation between this method and the existing clinical mass spectrometry methods was good. Conclusions:A candidate reference ICP-MS method for simultaneous determination of whole blood Fe and Cu was established. This is a simple and rapid method with good precision and spike recovery compared to the traditional one.

Analysis on the performance of aldosterone testing and the results of EQA in China / 中华检验医学杂志

Objective:To evaluate the performance of aldosterone testing in China through the External Quality Assessment (EQA) and improve the testing quality of aldosterone.Methods:Two kinds of EQA program for aldosterone were carried out in China, one of which is Routine EQA and the other is Trueness verification scheme. Lyophilized sera with 5 concentration levels were used as quality control of Routine EQA. The results were grouped according to the instrument. Target values and the coefficient of variation ( CV) were calculated in each group. Trueness verification scheme was verified by using frozen human sera of 3 concentration levels determined by the reference method, and the bias of each instrument group from the target value was calculated. Results:272 laboratories submitted the testing results, and 91.6% of laboratories used chemiluminescence method. The maximum CV was obtained by radioimmunoassay and liquid chromatography mass spectrometry, and the robust CVs were 14.6%-33.4% and 43.5%-53.9%, respectively. For chemiluminescence methods, the robust group CV was less than 10%. The results of the Trueness verification scheme showed that liquid chromatography mass spectrometry method was the most accurate method, with biases of -7.9%, 8.9% and -0.7% for the three quality controls. Diasorin system had the more accurate results deviated from the target by 58.7%, 7.9% and -2.1%, respectively. The results of other chemiluminescence methods were negatively correlated with the sample concentration, and one of them with a bias of 479%. Conclusions:The accuracy and comparability of aldosterone among laboratories in China are not satisfactory. Reagent manufacturers and laboratories should pay more attention to EQA, with the aldosterone results traceable to SI unit, and improve the test quality of aldosterone.

Detection and standardization of catecholamines and their metabolites / 中华检验医学杂志

Catecholamines include dopamine, norepinephrine and epinephrine. The main metabolites are 3-methoxytyramine, normetanephrine, metanephrine,homovanillic acid and vanillylmandelic acid. Detection of catecholamines and their metabolites is the cornerstone for the diagnosis of neuroendocrine tumors derived from neural crest such as pheochromocytoma, paraganglioma and neuroblastoma. Liquid chromatography tandem mass spectrometry has been widely used in the detection of catecholamines and their metabolites due to its high sensitivity and high specificity. However, the results of liquid chromatography tandem mass spectrometry in different laboratories are quite different and lack comparability. Accurate determination of catecholamines and their mtabolites in plasma and urine is currently a challenge in the field of clinical detectionbecause of their susceptibility to oxidative degradation, presence of numerous interferences and low concentration in plasma and urine samples.

Analysis of the oxidative products of DNA(8-oxo Gsn) and RNA(8-oxo dGsn) in patients with essential hypertension / 中华检验医学杂志

Objective:Analysis of the oxidative products of DNA and RNA in patients with hypertension by determination of 8-oxo Gsn and 8-oxo dGsn, respectively.Methods:This is an observational study. During August and December, 2018, 139 hypertension patients without other chronic diseases with an average age of (49.6±12.4) years old, and 139 apparently healthy volunteers without hypertension with an average age of (48.5±11.7) years old were recruited. Fasting morning urine were collected. The oxidative products of DNA: 8-oxo Gsn and the products of RNA: 8-oxo dGsn were analyzed using liquid chromatography tandem mass spectrometry. Urine Cr(U-Cr), and other serum biomarkers such as ALT, Cr, UA, Glu, TG, TC were analyzed using automatic biochemical analyzers.Results:of 8-oxo Gsn and 8-oxo dGsn were presented as median(quartile). Statistical analysis was performed using SPSS 22.0 and GraphPad Prism 5. Nonparametric test was used to compare the difference of 8-oxo Gsn and 8-oxo dGsn between the hypertension patients and the healthy controls. Results The DNA and RNA oxidative products of 8-oxo Gsn and 8-oxo dGsn in patients with hypertension and their U-Cr-corrected 8-oxo Gsn/U-Cr and 8-oxo dGsn/U-Cr were 14.38(10.39-19.91)ng/ml, 12.97(7.92-18.96)ng/ml, 1.10(0.88-1.38)μg/μmol and 0.96(0.75-1.30) μg/μmol, respectively, which were significantly higher than those in the healthy controls: 11.95(7.52-18.01) ng/ml, 10.12(6.42-15.04) ng/ml, 0.86(0.59-1.21) μg/μmol and 0.72(0.50-1.02) μg/μmol, respectively. After grouped by sex, 8-oxo Gsn in males, 8-oxo Gsn and 8-oxo dGsn in females showed no significant difference between patients with hypertension and healthy controls, however, after U-Cr correction, both males′ and females′ 8-oxo Gsn/U-Cr and 8-oxo dGsn/U-Cr in patients with hypertension were higher than that in the healthy controls.Conclusion:The oxidative products of DNA and RNA in patients with hypertension were significantly higher than that in healthy controls.

Standardization of aldosterone and harmonization of renin in China / 中华检验医学杂志

Aldosterone and renin are important clinical indicators in the diagnosis and treatment of hypertension. There are two kinds of methods for aldosterone detection, including immunoassay and mass spectrometry. The coefficients of variation within the methods, as well as the deviation between methods are large.And the results of immunoassay are much higher than that of mass spectrometry. Clinically, renin tests mainly include renin concentration and renin activity, both of which have advantages and disadvantages in the diagnosis and treatment of hypertension. The disordered test results of aldosterone and renin brings great trouble to the clinical diagnosis and treatment. Standardization/consistency is urgently needed to make the test results of each laboratory more accurate and comparable. The standardization project for renin and aldosterone in China will greatly promote the standardization process in order to provide accurate and comparable clinical tests.

Commutability assessment of 16 processed materials for 17-hydroxyprogesterone based on two approaches in China / 中华检验医学杂志

Objective:The aim of this study is to evaluate the commutability of 16 processed materials for 17-hydroxyprogesterone by using 2 commutability assessment approaches.Methods:52 serum specimens were collected in Clinical Laboratory Department of Beijing Hospital from February 2018 to June 2019. According to the report of the Clinical and Laboratory Standards Institute (EP14-A3) document and the recommendations of the International Federation of Clinical Chemistry and Laboratory Medicine (IFCC) working group on commutabilityassessment, serum 17-hydroxyprogesterone isotope diluent chromatogram tandem mass spectrometry (ID-LC/MS/MS) was used for comparison. Three clinical routine analysis systems (1 radioimmunoassay, 2 LC/MS analysis methods) were used to determine the concentration of 17-hydroxyprogesterone in 52 human serum samples and 16 processed materialsfor commutabilityassessment.Results:Combined with the results of the two commutability assessment, all accuracy verification materials and national steroid hormone standards showed good commutability in the LC/MS analysis system, and 6/9 EQA materials showed commutability in the three routine analysis systems.All materials showed good commutability in the LC/MS analysis system of bias difference method.Conclusions:The two kinds of commutability assessment results are different. Bias difference method has more clinical value, but it has certain application limitations. The use of fresh frozen human serum as a quality assessment materialfor serum 17-hydroxyprogesterone is meets the commutability requirement.

Comparison of Six Automated Immunoassays With Isotope-Diluted Liquid Chromatography-Tandem Mass Spectrometry for Total Thyroxine Measurement

BACKGROUND: Accurate serum total thyroxine (TT4) measurement is important for thyroid disorder diagnosis and management. We compared the performance of six automated immunoassays with that of isotope-diluted liquid chromatography-tandem mass spectrometry (ID-LC-MS/MS) as the reference method. We also evaluated the correlation of thyroid stimulating hormone (TSH) with TT4 measured by ID-LC-MS/MS and immunoassays. METHODS: Serum was collected from 156 patients between October 2015 and January 2016. TT4 was measured by immunoassays from Abbott (Architect), Siemens (ADVIA Centaur XP), Roche (E601), Beckman-Coulter (Dxi800), Autobio (Autolumo A2000), and Mindray (CL-1000i), and by ID-LC-MS/MS. Results were analyzed using Passing-Bablok regression and Bland-Altman plots. Minimum requirements based on biological variation were as follows: a mean bias of ≤4.5% and total imprecision (CV) of ≤3.7%. RESULTS: All immunoassays showed a correlation >0.945 with ID-LC-MS/MS; however, the slope of the Passing-Bablok regression line varied from 0.886 (Mindray) to 1.23 (Siemens) and the intercept from −12.8 (Siemens) to 4.61 (Mindray). Only Autobio, Beckman-Coulter, and Roche included the value of one in the 95% confidence interval for slope. The mean bias ranged from −10.8% (Abbott) to 9.0% (Siemens), with the lowest value noted for Roche (3.5%) and the highest for Abbott (−10.8%). Only Abbott and Roche showed within-run and total CV ≤3.7%. CONCLUSIONS: Though all immunoassays correlated strongly with ID-LC-MS/MS, most did not meet the minimum clinical requirement. Laboratories and immunoassay manufacturers must be aware of these limitations.

Determination of serum high- and low-density lipoprotein cholesterol by ultracentrifugation-high performance liquid chromatography--improvement and verification of β-quantification method / 中华检验医学杂志

Objective To propose and validate a reduced volume β-quantification method to measure serum high-density lipoprotein cholesterol (HDL-C) and low-density lipoprotein cholesterol (LDL-C). Methods The reduced volume β-quantification method involved separation of LDL and HDL by ultracentrifugation and preparation of HDL by chemical precipitation. The sampling and reconstitution of the bottom fractions were performed gravimetrically and sample volume was thus decreased from 5 to 0.8 ml. High performance liquid chromatography was used to determine the cholesterol concentration of bottom fractions and HDL-C in the supernatant. Serum levels of LDL-C depended on a calculation of bottom fractions cholesterol minus HDL-C. Results The total CVs for HDL-C and LDL-C were 0.65％ -1.75％ and 0.63％ -1.11％. The results of the developed method were consistent with the current reference method and well within the allowable bias for Cholesterol Reference Method Laboratory Network surveys. Conclusion A new method for the measurement of HDL-C and LDL-C has been established. This method requires a small amount of serum and is easy to operate, exhibiting a desirable precision and accuracy. It is reliable and can be used as a candidate reference method for HDL-C and LDL-C.

Comparison of ID-LC-MS/MS and two CLIAs in measuring plasma aldosterone / 中华检验医学杂志

Objective Accurate measurement of aldosterone is critical in the diagnosis of primary aldosteronism. We compared the harmonization of three assays including isotope dilution liquid chromatography-tandem mass spectrometry (ID-LC-MS/MS) and two chemiluminescence immunoassays (CLIAs:system A and system B) for the aldosterone measurement. Methods A total of 45 plasma samples, 4 quality control materials, 5 lyophilized bovine serums, and 3 fresh frozen human serum pools were measured by three assays respectively. Based on CLSI EP15-A3 rule, the precision was assessed by coefficient of variance. Deming regression and Bland&Altman plots was performed for method comparison, and correlation coefficient was calculated for concordance (CCC). Results All three methods met the performance criteria based on desirable biological variation for precision (<7.35％). System A showed a relevantly good correlation and comparability with ID-LC/MS/MS (R2=0.985, CCC=0.967), while System B showed relevantly bad correlations and comparability with both System A (R2=0.538, CCC=0.605) and ID-LC/MS/MS (R2=0.547, CCC=0.528).. However, the average relevant bias of two CLIAs exceeded the bias requirement derived from biological variation (18.60％). Conclusion Significant differences were found in the measurement of plasma aldosterone using ID-LC-MS/MS and two CLIAs, which urges the establishment of traceability hierarchy and improvement of reagents' specificity for standardization of aldosterone measurement in clinical settings.

Comparison between the triglycerides determination by HPLC in China and reference procedure used in the isotope dilution mass spectrometry CDC CRMLN / 中华检验医学杂志

Objective To evaluate the comparability and consistency of two kinds of triglycerides reference methods, one of which is the isotope dilution liquid chromatography-mass spectrometry (LC/MS) in the Cholesterol Reference Method Laboratory Network (CRMLN), the other isthehigh-performance liquid chromatography (HPLC) method for triglyceride detection in China. Methods 52 fresh frozen sera with triglycerides levels among 0.45-4.52 mmol/L were determined by LC/MS and HPLC. After evaluation the precision and accuracy of the two methods,a series of analyses were conducted including plotting to scatter plots and deviation graphs, testing outliers, selecting the best fitting regression models and calculating the regression equations and parameters, and so on. The expected deviation at the level of medical decision of triglycerides and the 95％confidence range were statistically analyzed.Results For HPLC method, the CV of instrument measurement was 0.29％(0％-1.16％), the total CV of samples measurement was 0.54％(0.04％-1.28％), and the average bias of the reference materials was 0.22％(-0.43％-0.68％). ForLC/MSmethod,the CV of instrument measurement was 0.55％(0％-1.68％),the total CV of samples measurement was 0.79％(0％-1.93％), and the average bias of the NIST reference materials was 0.09％(-0.73％-1.29％). No outlier was found from the scatter plots and the statistical analysis and the linear regression was fitted to analyze the results of the two methods. The linear regression parameters of two methods for 52 fresh frozen human sera were as follows:the slope was 0.9988,the standard error of slope was 0.0035, the intercept was 0.0037mmol/L, the standard error of intercept was 0.0030 mmol/L, the standard error of Y-estimate was 0.0236 mmol/L,and the correlation coefficient was 0.9997. Compared with the LC/MS method,the absolute deviation of fresh sera by HPLC method was-0.001 mmol/L (-0.070-0.056 mmol/L), with a relative deviation of 0.13％ (-2.01-2.83％). T-test showed no statistically significant difference between the two methods. The expected deviations at the triglycerides medicine decision level were within the 95％confidence range,and the expected deviations were far less than the allowable error. Conclusions The HPLC method of triglyceridesdetetion has good consistency and comparability with LC/MS method as one of the reference methods of CRMLN. Because of the advantages of HPLC method such as low cost, simplicity,less technical need,and better precision,HPLC method is expected to play an important role in the process of standardization and traceability of serum triglycerides.