Table of Content

30 March 2014, Volume 29 Issue 3

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Orginal Article

Problems and thinking of reagents for clinical molecular diagnosis in China

LI Jinming.

2014, 29(3):  199-201.  DOI: 10.3969/j.issn.1673-8640.2014.03.001

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The methods used in commercial reagents for clinical molecular diagnosis have experienced the development courses of molecular hybridization,polymerase chain reaction (PCR)-agarose gel electrophoresis,PCR-enzyme-linked immunosorbent assay (ELISA) and real-time PCR. The pursuit of simple operation influences on the repeatability and sensitivity of the reagents. Recently,molecular diagnosis reagents based on different technique platforms for personalized medicine,including PCR-Luminex,PCR-sequencing,high resolution melting curve analysis and so on,have entered the clinical laboratories. During current and future quite a long time,manual operation in routine work of clinical molecular diagnosis laboratories will still be the major way in China. Also,home-brew reagents should be used in clinical laboratories. How to develop molecular diagnosis reagents that be suitable for laboratories in medical establishment of basic unit is worth to ponder,however,there is no doubt that the automatic system for routine testing is the development direction of clinical molecular diagnosis.

Evolutions in half a century: the course of change and development of current molecular diagnostics techniques

WU Zhiyuan, ZHANG Chen, GUAN Ming.

2014, 29(3):  202-208.  DOI: 10.3969/j.issn.1673-8640.2014.03.002

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Molecular diagnostics techniques uses molecular biology techniques to analyze the nucleotide acid biomarkers of humen or pathogens,and to assist the clinical diagnosis of diseases. Since the innovation of liquid hybridization,molecular diagnostics has witness its rapid development for over half a century. Here we will assort the common molecular diagnostics procedures according to their methodological origins,so as to review the evolution of current clinical techniques,and to get a technical perspective on the development of molecular diagnostics.

Development of warfarin-related pharmacogenomics and its prospect of clinical application

WU Jiong, WU Shengchao, GUO Wei, PAN Baishen.

2014, 29(3):  209-214.  DOI: 10.3969/j.issn.1673-8640.2014.03.003

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Warfarin,a commonly prescribed anticoagulant,has warried lots of patients and doctors because of its narrow therapeutic range and large interindividual variability in daily dose. During recent years,many studies focus on genetic polymorphisms,regarding it as one of the main effects which lead to dose difference between individuals. Some of them established and verified pharmacogenetic-based warfarin-dosing algorithms and evaluated the clinical significance. Our review exhibits the latest development of warfarin-related pharmacogenomics home and abroad,in order to provide references for further clinical studies.

Improvement on the detection rate of Mycobacterium tuberculosis in fluorescence quantitation PCR by increasing sputum volume and modifying ultrasonic extraction procedure

ZHANG Jun, WANG Xiaofei, WANG Ruidong, HAN Min, JING Lingjie, CHEN Jin.

2014, 29(3):  215-218.  DOI: 10.3969/j.issn.1673-8640.2014.03.004

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Objective　To increase the detection rate of Mycobacterium tuberculosis(MTB) in real-time fluorescence quantitation polymerase chain reaction (PCR) by increasing sputum volume and modifying ultrasonic extraction of MTB DNA. Methods　Sputum samples from 206 tuberculosis patients and 103 non-tuberculosis patients as controls were analyzed. Routine sputum MTB smear and liquid culture of MTB by American BD MAGIT960 system were performed. At least 5mL sputum was collected from each patient,and 2-3-fold volume of 4% NaOH solution was added to the sputum. After liquefaction,1 mL solution was used for the routine DNA extraction procedure,and the remaining part was used for the modified ultrasonic extraction procedure. Both of the extracted DNA were quantitated by real-time fluorescence quantitation PCR. Results　By the modified ultrasonic extraction procedure,the MTB DNA positive detection rate increased from 87.5%(confidence interval 81.4%-93.6%)to 95.5% (confidence interval 91.7%-99.4%)(P<0.05)in the smear positive and culture positive tuberculosis patients,and the positive detection rate increased from 57.4%(confidence interval 47.5%-67.4%) to 83%(confidence interval 75.4%-90.6%) in the smear negative and culture positive tuberculosis patients (P<0.01). The quantity results of increasing sputum volume and modifying ultrasonic extraction procedure increased 14 folds in average by the modified ultrasonic extraction procedure. Conclusions　The increasing sputum volume and modifying ultrasonic extraction procedure increase the positive detection rate of real-time fluorescence quantitation PCR for tuberculosis,which can be recommended for routine laboratory use.

Correlationship research of ALOX12 gene polymorphism with type 2 diabetes mellitus and diabetic nephropathy

LI Qi,WANG Yan, QIU Yifan, DONG Ling, CHEN Kelin

2014, 29(3):  219-225.  DOI: 10.3969/j.issn.1673-8640.2014.03.005

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Objective　To investigate the correlation of a single nucleotide polymorphism (SNP) locus rs1126667 (R261Q) in the arachidonate 12-lipoxygenase (ALOX12) gene with type 2 diabetes mellitus(T2DM) and diabetic nephropathy(DN). Methods　The SNP of ALOX12 gene was studied using a case-control method by single base extension reactions and matrix-assisted laser desorption ionization-time of flight mass spectrometry platform. A case-control study enrolled 223 T2DM patients [DN group (134 patients) and T2DM without nephropathy group (89 patients)] and 120 healthy subjects (healthy control group). Results　ALOX12 gene locus rs1126667 was consistent with Hardy-Weinberg equilibrium law,and the frequencies of ALOX12 gene locus rs1126667 G/G,G/A and A/A genotypes in the healthy control group were 26%,47% and 27%. The frequencies in T2DM group were 30%,48% and the 22%,and odd ratios (OR) were 1,0.8 and 0.76. The frequencies in DN group were 31%,45% and 24%,and OR were 1,0.88 and 0.94. The frequencies in T2DM without nephropathy group were 28%,54% and 18%,and OR were 1,1.06 and 0.61. No statistical significance in all groups was found(P>0.05). A allele frequencies in the healthy control group,T2DM group,DN group and T2DM without nephropathy group were 50%,46%,47% and 45% without statistical significance(P>0.05). Conclusions　No correlation of the ALOX12 gene polymorphism with T2DM or DM is found in the northeast Han Chinese.

Detection of BCR-ABL fusion gene by multiplex RT-PCR combining with capillary electrophoresis

Lü Xiaonan，YE Huiming，GU Long，ZENG Jimeng

2014, 29(3):  226-232.  DOI: 10.3969/j.issn.1673-8640.2014.03.006

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Objective　To establish a method for detecting BCR-ABL fusion gene by multiplex reverse transcription polymerase chain reaction ( RT-PCR),and to provide a useful tool for chronic myeloid leukemia (CML) auxiliary diagnosis. Methods　The construction of BCR-ABL fusion gene as positive template was made by overlap extension method. The method of multiplex RT-PCR combining with capillary electrophoresis was designed and optimized. Its primary performance was evaluated. Results　The lower detection limit for BCR-ABL fusion gene (ela2,e13a2 and e14a2) by multiplex RT-PCR was 10²-10³ copies/μL. The positive rate of this method to detect 50 CML patients was 86.0%(e13a2: 20.0% and e14a2: 66.0%). Compared with the results of chromosome karyotype analysis,the positive coincidence rate was 97.6%,and the negative coincidence rate was 75.0%. The 2 methods have high consistency (Kappa=0.765,P>0.05). Conclusions　The method of multiplex RT-PCR combining with capillary electrophoresis to detect BCR-ABL fusion gene is simple and rapid with good specificity and sensitivity and is very suitable for the clinical detection of CML.

Analysis on the results of chromosomal microarray in patients with chromosome abnormality

YAO Ruen，FU Qihua，YU Yongguo

2014, 29(3):  233-236.  DOI: 10.3969/j.issn.1673-8640.2014.03.007

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Objective　To analyze the results of patients with chromosome abnormality by chromosomal microarray,and validate and depict patient's karyotype. Methods　Genotyping array was applied to assess in 9 patients with hypophrenia and deformity. The samples with karyotype analysis abnormality were determined. Comparison between karyotyping and microarray analysis was considered to validate the clinical utility of chromosomal microarray analysis. Results　Two Turner syndrome patients had completely concordant results from karyotype analysis and chromosomal microarray. Three patients showed abnormal karyotyping results (2 patients with excess trabant and 1 patients with chromosomal inversion) were detected negative with chromosomal microarray. Four patients with different size and location of deletions and duplication were also detected abnormal with chromosome microarray,but with significant deviation on location. Conclusions　Compared with chromosomal microarray analysis,karyotype analysis has low accuracy in diagnosing patients with hypophrenia and deformity. The abilities of locating and reckoning chromosomal deletion and duplication by chromosomal microarray are insufficient for clinical diagnostic purpose,but karyotype analysis is still efficient in detecting balanced changes in chromosome.

Analysis on mutational spectrum of GATA5 gene associated with bicuspid aortic valve

XU Lei，YUAN Fang，LI Ruogu，WANG Qian，ZHENG Hongzhen，YANG Yiqing

2014, 29(3):  237-240.  DOI: 10.3969/j.issn.1673-8640.2014.03.008

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Objective　To analyze the mutational spectrum of GATA5 gene associated with congenital bicuspid aortic valve(BAV). Methods　A total of 150 patients with congenital BAV and a total of 200 healthy subjects as controls were enrolled. The clinical data were collected,and the peripheral venous blood specimens were prepared. The genomic DNA was isolated by DNA purification kit. The coding regions and splice junction sites of GATA5 gene were amplified by polymerase chain reaction,and the amplicons were sequenced by di-deoxynucleotide chain termination technique. The acquired sequences were aligned with those of GATA5 released from GenBank to identify the likely sequence mutations. The online software MUSCLE and MutationTaster were used to evaluate whether the altered amino acids were conserved evolutionarily and whether the mutations were causative,respectively. Results　Two novel heterozygous GATA5 mutations,p.M219I and p.T289I,were identified in 2 congenital BAV patients,respectively,with a mutational prevalence of approximately 1.33%. The 2 mutations,which altered the amino acids completely conserved evolutionarily,were absent in the 200 controls and were both predicted to be pathogenic. Conclusions　This study links novel GATA5 mutations to congenital BAV,which helps to reveal new molecular mechanism underlying congenital BAV.

Investigation on the evaluation of the uncertainty of hepatitis B virus DNA determination by fluorescence quantitative PCR

JIANG Lingli, WANG Xueliang, XIAO Yanqun, WANG Hualiang.

2014, 29(3):  241-244.  DOI: 10.3969/j.issn.1673-8640.2014.03.009

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Objective 　To investigate the uncertainty of hepatitis B virus (HBV) DNA determination by fluorescence quantitative polymerase chain reaction (PCR) according to quality control data,and evaluate the feasibility. Methods　The uncertainty of Type A was evaluated by the data of within-run and between-run coefficients of variation. The uncertainty of Type B was evaluated according to the data of external quality assessments from the National Center for Clinical Laboratory and College of American Pathologists(CAP). The combined uncertainties and expanded uncertainties were evaluated and analyzed. Results　The expanded uncertainties in 10⁴-10⁵ IU/mL and 10⁶-10⁷ IU/mL concentrations were 0.521 6 and 0.529 8(k=1.96,n=1),according to the data from the National Center for Clinical Laboratory. The expanded uncertainties in 10⁴-10⁵ IU/mL and 10⁶-10⁷ IU/mL concentrations were 0.973 1 and 0.977 5(k=1.96,n=1),according to the data from CAP. Conclusions　It is not bilievable to evaluate the uncertainty of HBV DNA determination by fluorescence quantitative PCR,and it is suitable to evaluate the uncertainty using the results traced back to the national standard substance.

Analysis on clinical distribution and resistance characteristic of Acinetobacter baumannii isolated clinically from 2008-2012

XU Yiming，WANG Bei，JIANG Xiaofei.

2014, 29(3):  245-248.  DOI: 10.3969/j.issn.1673-8640.2014.03.010

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Objective　To investigate the clinical distribution and resistance characteristic trend of Acinetobacter baumannii in Huashan Hospital isolated clinically from January 2008 to September 2012,and to provide the reference for clinical rational using of antimicrobial agents. Methods　The drug-resistance of Acinetobacter baumannii and resistance characteristic trend to common antimicrobial agents were analyzed by disk agar diffusion method. Results　In the past 5 years,the detection rate of clinical isolated Acinetobacter baumannii from sputum specimens was highest,and mainly distributed in neurosurgery department and intensive care unit. The isolation rate in neurosurgery department presented obvious increasing trend from 19.6% in 2008 to 35.3% in 2012. The resistance of clinical isolated Acinetobacter baumannii was serious,and carbapenem antimicrobial agents had the obvious increasing trend. The resistance rate of imipenem rase from 57.5% in 2008 to 71.2% in 2012,the resistance rate of meropenem rase from 56.8% in 2008 to 71.7% in 2012,and the resistance rates of other lactams,cephalosporins,aminoglycosides and fluoroquinolones were 70.0%-80.0%. Conclusions　The infection of Acinetobacter baumannii is serious in mainly neurosurgery department and intensive care unit. The clinical isolates are commonly resistant to all kinds of antimicrobial agents,and carbapenem antimicrobial agents present a obvious increasing trend. Monitoring the resistance of Acinetobacter baumannii should be strengthened for preventing resistant bacteria from spreading.

Plasma lipoprotein-associated phospholipase A2 level and its clinical significance in patients with stroke

YE Yun，LI Suliang

2014, 29(3):  249-253.  DOI: 10.3969/j.issn.1673-8640.2014.03.012

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Objective　To investigate the changes of lipoprotein-associated phospholipase A2 (Lp-PLA2) level in patients with stroke,and to research the correlation with the volume of cerebral infarction and the severity of neurological impairment in patients with ischemic stroke. Methods　A total of 180 ischemic stroke patients,165 hemorrhagic stroke patients and 105 healthy subjects (healthy control group) were enrolled. By enzyme-linked immunosorbent assay (ELISA),the plasma Lp-PLA2 levels were determined,and the receiver operating characteristic (ROC) curve was used to evaluate the efficacy of Lp-PLA2 in the diagnosis of ischemic stroke and hemorrhagic stroke. The blood lipids [total cholesterol (TC),triglyceride (TG),high-density lipoprotein cholesterol (HDL-C) and low-density lipoprotein cholesterol (LDL-C)],glucose (Glu) and fibrinogen (FIB) were determined. The volumes of cerebral infarction were measured by cranium magnetic resonance imaging,and the severities of neurological impairment were assessed by the National Institutes of Health Stroke Scale.The correlation of plasma Lp-PLA2 with the National Institutes of Health Stroke Scale was analyzed. Results　The levels of Lp-PLA2,TC,HDL-C,LDL-C and FIB among ischemic stroke,hemorrhagic stroke and healthy control groups had statistical significance (P<0.05). TG and Glu had no statistical significance between ischemic stroke and hemorrhagic stroke groups (P=0.133 and 0.067),and there were statistical significance with those in healthy control group(P<0.05). The area under the ROC curve (AUC) was 0.905,and the optimal cut-off value was 42.35 μg/L. The diagnostic sensitivity and specificity of plasma Lp-PLA2 were 81.5% and 80.0%. Plasma Lp-PLA2 increased with the increase of the volume of cerebral infarction,but it had no statistical significance(H=0.372,P=0.719).Spearman correlation analysis showed that plasma Lp-PLA2 had significant correlation with the National Institutes of Health Stroke Scale(P<0.05). Conclusions　Plasma Lp-PLA2 increases significantly in patients with ischemic stroke,and it is significantly correlated with the National Institutes of Health Stroke Scale. Plasma Lp-PLA2 is an independent risk factor for ischemic stroke,and it may be a prediction index of ischemic stroke and a useful parameter for the severity of patient's condition.

Investigation on the detection and its significance of vascular risk factors in the subjects with different blood pressure

WANG Guoqiang, YANG Cailing, GUO Lijuan

2014, 29(3):  254-257.  DOI: 10.3969/j.issn.1673-8640.2014.03.013

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Objective　To detect the vascular risk factors in the subjects with different blood pressure，in order to investigate the significance of vascular risk factors in the diagnosis and prevention of cardiovascular diseases. Methods　A total of 246 subjects were classified into normal blood pressure group（35 cases），normal blood pressure high value group（46 cases），grade 1 hypertension group（52 cases），grade 2 hypertension group（55 cases） and grade 3 hypertension group（58 cases） according to the Guidelines for the Prevention and Treatment of Hypertension in China 2010. Serum endothelin-1（ET-1），high-sensitivity C-reactive protein（hs-CRP） and homocysteine（Hcy） were determined，and the carotid artery intima-media thickness（IMT） was measured by color ultrasound. The difference among the groups was analyzed. Results　The Hcy levels of grade 2 and 3 hypertension groups were higher than those of the other 3 groups（P＜0.05），and the Hcy level of grade 3 hypertension group was higher than that of grade 2 hypertension group（P＜0.05）. The hs-CRP level of grade 3 hypertension group was higher than those of the other 4 groups（P＜0.05），the hs-CRP level of grade 2 hypertension group was higher than those of normal blood pressure and normal blood pressure high value groups（P＜0.05），and the hs-CRP level of grade 1 hypertension group was higher than that of normal blood pressure group（P＜0.05）. The ET-1 levels of grade 1，2 and 3 hypertension groups were higher than those of normal blood pressure and normal blood pressure high value groups（P＜0.05），and the ET-1 levels of grade 1，2 and 3 hypertension groups were significant statistically（P＜0.05）. The IMT detection results of grade 3 hypertension group were higher than those of the other 4 groups（P＜0. 05），and those of grade 2 hypertension group were higher than those of normal blood pressure and normal blood pressure high value groups（P＜0.05）. ET-1，hs-CRP，Hcy and IMT increased with systolic pressure，and the correlation coefficients（r） were 0.302，0.214，0.192 and 0.285（P＜0.05）； ET-1，hs-CRP and Hcy had positive correlations with IMT（r=0.358，0.249 and 0.226，P＜0.05）. Conclusions　There are good correlations among ET-1，hs-CRP and Hcy with IMT and corresponding blood pressure，and the ET-1 is the most sensitive in the progress of blood pressure change. ET-1，hs-CRP，Hcy and IMT should be used as routine physical examination parameters in order to prevent or stop cardiovascular risk. Especially，the subjects with normal blood pressure at present should be paid attention.

The influence of acrosome enzyme and hyaluronidase in human sperm acrosome on male fertility

MA Xiaoping, GAO Xiaoqin, YANG Yanping, YANG Zhe

2014, 29(3):  258-261.  DOI: 10.3969/j.issn.1673-8640.2014.03.014

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Objective　To investigate the influence of enzymes in human sperm acrosome on male fertility. Methods　According to the World Health Organization standard method,52 cases were enrolled as infertile group,and 12 cases were enrolled as normal control group[sperm density (50-100)×10⁹/L and sperm motility 60%-90%],and semen routine analysis was performed. According to the density of sperm,the infertile group was classified into infertile group B1(<20×10⁹/L),C1(20×10⁹ -40×10⁹/L) and D1(>40×10⁹/L),and according to the motility of sperm,the infertile group was classified into infertile group B2(<30%),C2(30%-50%) and D2(>50%). The positive reaction rate and activity intensity of acrosome enzyme (ACE) were detected by fixed gelatin substrate film technique . The positive reaction rate and activity intensity of hyaluronidase (HYD) were detected by modified sodium hyaluronate-gelatin membrane. Results　There was a significant correlation between the positive reaction rate and activity intensity of ACE with sperm density,sperm motility and sperm deformity rate [correlation coefficients (r) were 0.797,0.867; 0.831,0.860 and -0.625,-0.546,P<0.01]. There was a significant correlation between the positive reaction rate and activity intensity of HYD with sperm density,sperm motility and sperm deformity rate (r were 0.832,0.855; 0.842,0.830 and -0.625,-0.554,P<0.01). The sperm deformity rate in infertile group was higher than that in normal control group (P<0.01). The positive reaction rates and activity intensities of ACE and HYD in infertile group C1,D1 and C2,D2 were significantly lower than those in normal control group (P<0.01). The activity of enzymes increased with the sperm density and motility. Conclusions　The detections of sperm ACE and HYD are effective parameters for the analysis of male fertility.

The accuracy evaluation of nucleated red blood cell count determined by SYSMEX XN-1000 automatic hematology analyzer

WANG Yefei, ZHOU Yi, DING Lei, XIA Wenquan, HU Yiqun.

2014, 29(3):  262-265.  DOI: 10.3969/j.issn.1673-8640.2014.03.015

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Objective　To investigate the accuracy of nucleated red blood cell(NRBC) count determined by SYSMEX XN-1000 automatic hematology analyzer(XN-1000). Methods　A total of 66 peripheral blood samples were collected and determined for NRBC by XN-1000,and the results of NRBC% were compared with those by flow cytometry (FCM) and manual microscopy for the same sample. The manual microscopy was as golden standard. Results　There was a good correlation for NRBC between XN-1000 and microscopy(r=0.927),but the correlation was lower than that between FCM and manual microscopy (r=0.956).The correlation between XN-1000 and FCM was the best (r=0.994). The results of XN-1000 and FCM with manual microscopy had no statistical significance(P>0.05). XN-1000 for NRBC% had a better specificity (95.7%) than FCM. Conclusions　XN-1000 automatic hematology analyzer has good performance for detecting NRBC in peripheral blood rapidly,accurately and effectively.

Investigation on the significance of NEUT-X and NEUT-Y parameters for the diagnosis of bacterial infection

CUI Ling，LIU Yun，JIN Ligang，BAI Xiaosong

2014, 29(3):  266-269.  DOI: 10.3969/j.issn.1673-8640.2014.03.016

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Objective　To investigate the clinical significance of Sysmex XT-2000i automatic hematology analyzer's neutrophil parameter NEUT-X and NEUT-Y in the diagnosis of bacterial infection. Methods　Blood samples were collected from 114 patients with bacterial infection,96 patients with bacterial culture negative and 142 healthy subjects(healthy control group) to detect and analyze NEUT-X,NEUT-Y,C-reactive protein (CRP) and toxity indices (TI). Results　NEUT-X and NEUT-Y were 1 391.4±42.3 and 450.9±40.9 respectively in bacterial infection group,which were significantly higher than in bacterial culture negative group (1 344.5±37.0 and 422.7±45.9) and healthy control group(1 351.7±33.7 and 415.8±32.2) with statistical significance(P<0.01). NEUT-X and NEUT-Y were correlated with CRP and TI. For the diagnosis of bacterial infection,CRP had the best diagnosis efficacy,and NEUT-X had the secondary diagnosis efficacy. When the cut-off value of CRP and NEUT-X were 20.5 and 1 370.5,the sensitivities were 94.7% and 71.4%,and the specificities were 71.4% and 75.6%,respectively. Conclusions　NEUT-X and NEUT-Y parameters increase significantly after bacterial infection. It can be used as assistant diagnosis parameters of bacterial infection.

Application of serum abnormal prothrombin in clinical diagnosis of primary hepatocellular carcinoma

PU Juebiao,WANG Xuefeng,PENG Yibing

2014, 29(3):  270-273.  DOI: 10.3969/j.issn.1673-8640.2014.03.017

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Objective　To investigate the significance of serum abnormal prothrombin ［protein induced by vitamin K absence or antagonist Ⅱ(PIVKA Ⅱ)］ in clinical diagnosis of primary hepatocellular carcinoma. Methods　There were 365 samples from inpatients in Ruijin Hospital, including 100 patients with primary hepatocellular carcinoma and the other 265 patients with no primary hepatocellular carcinoma (59 cases of chronic liver disease, 50 cases of gastrointestinal cancer, 50 cases of secondary liver cancer, 56 cases of other hepatopathy and 50 cases of healthy controls). Serum alpha fetoprotein (AFP) and PIVKA Ⅱlevels were detected by Roche Cobas e601 automatic immunity analyzer and LUMIPULSE G1200 automatic immunity analyzer, respectively. Data were analyzed statistically by SPSS 16.0 software. Results　Serum AFP and PIVKA Ⅱlevels were significantly higher in primary hepatocellular carcinoma group than those in other disease group and healthy controls. Serum AFP had a sensitivity of 63.00% and a specificity of 84.91% in the diagnosis of primary hepatocellular carcinoma, while PIVKA Ⅱhad a sensitivity of 74.00% and a specificity of 89.81%. The results of receiver operating characteristic (ROC) curve showed that the areas under the curve of AFP and PIVKA Ⅱwere 0.789 and 0.873, respectively. The diagnosis sensitivity and specificity of the combination determination of AFP and PIVKA Ⅱreached to 81.00% and 98.49%. Conclusions　Serum PIVKA Ⅱ has a better diagnosis significance than AFP, and can be a tumor marker in the diagnosis of primary hepatocellular carcinoma. Moreover, the combination determination of AFP and PIVKA Ⅱcan improve the diagnosis efficiency for clinical primary hepatocellular carcinoma.

The cytokine changes of Th1/Th2 in patients with acute-on-chronic hepatitis B liver failure and their relationships with liver function parameters

GUO Yonghong, ZHOU Yun, ZHANG Ying, HE Yu, JI Guangxi, MA Zhiyuan, CHENG Cheng, FAN Chao, ZHANG Lingyun, JIA Zhansheng

2014, 29(3):  274-278.  DOI: 10.3969/j.issn.1673-8640.2014.03.018

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Objective　To investigate the cytokine levels of T helper cell 1/2 (Th1/Th2) detected by cytometric bead array (CBA) in patients with acute-on-chronic hepatitis B liver failure (ACHBLF),and their change significance in ACHBLF. Methods　The Th1/Th2 cytokine levels in 33 patients with ACHBLF and 20 patients with chronic hepatitis B (CHB) were detected by CBA. A total of 15 healthy subjects were enrolled as controls. The serum interferon gamma (IFN),interleukin 2 (IL-2),interleukin 4 (IL-4),interleukin 6 (IL-6),interleukin 10 (IL-10) and tumor necrosis factor alpha (TNF-α) levels were detected,and the changes of Th1/Th2 cytokine levels were analyzed. Results　The serum TNF-α,IFN-γ,IL-4 and IL-6 levels of ACHBLF group were significantly higher than those of CHB and control groups (P<0.05).The serum IL-2 and IL-10 had no statistical significance between ACHBLF and CHB groups and between ACHBLF and control groups (P>0.05) . There was no significant correlation of IL-4,IL-6,TNF-α in ACHBLF group with total bilirubin (TBil),albumin (Alb) and prothrombin activity (PTA). Conclusions　CBA can determine a variety of cytokines simultaneously and is suitable for the clinical application. The changes of TNF-α,IFN-γ,IL-4 and IL-6 will involve to the pathogenesis and development of ACHBLF. There is not found the relationship between cytokines and liver function.

Research on the expressions of KAI1 gene, TIMP-1 and MMP-9 and their correlations with clinicopathologic features of non-small cell lung cancer

ZHU Bo, ZHAO Huiliu, WANG Ying, LAO Ming, HUANG Zhaodong, LIU Zhimin, XIE Juan, OU Chao.

2014, 29(3):  279-282.  DOI: 10.3969/j.issn.1673-8640.2014.03.019

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Objective　To investigate the expressions of KAI1 gene,tissue inhibitor 1 of metalloproteinase (TIMP-1) and matrix metalloproteinase 9 (MMP-9) in non-small cell lung cancer (NSCLC) and their correlations with clinicopathologic features of NSCLC. Methods　The expressions of KAI1,TIMP-1 and MMP-9 in 73 specimens of NSCLC tissues,51 specimens of corresponding adjacent tissues and 13 specimens of non-cancerous lung tissue were determined by immunohistochemistry. Their correlations with clinicopathologic features were analyzed by χ² test and Pearson analysis. Results　In 73 specimens of NSCLC tissues and 64 cases of corresponding adjacent and non-cancerous lung tissues,the positive rates of KAI1 and TIMP-1 were 39.7%,74.0% and 90.6%,37.5% with statistical significance (P<0.05),and the positive rates of MMP-9 were 79.4%,67.1% without statistical significance (P>0.05). The expression of KAI1 was correlated to TNM stage,lymph node metastasis,histologic subtype and differentiation with statistical significance (P<0.05),and the TIMP-1 and MMP-9 were correlated to TNM stage and lymph node metastasis (P<0.05),but they were not correlated to age and sex. The expression of KAI1 was negatively correlated with the expressions of TIMP-1 and MMP-9 (r=-0.463 7,-0.344 5,P<0.05). Conclusions　The KAI1,TIMP-1 and MMP-9 are related to the development of lung cancer. The combination determination of them will be the important prognosis reference indicator for the occurrence and development of NSCLS.

Methodological study of serum free fatty acids in patients with coronary disease based on derivation reaction combined with gas chromatography-mass spectrometry

LI Qiang, WANG Shijie, HU Xiaoying, WEI Yangfei, QI Huanyang

2014, 29(3):  283-287.  DOI: 10.3969/j.issn.1673-8640.2014.03.020

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Objective　To establish a method for determining the serum concentrations of 11 free fatty acids(FFA) in patients with coronary disease. Methods　The serum concentrations of 11 FFA in patients with coronary disease were determined by gas chromatography-mass spectrometry(GC/MS) combined with derivation reaction and N,O-bis(trimethylsilyl) trifluoroacetamide(BSTFA) as derivative reagent. Results　Chromatographic peaks of 11 FFA were separated completely,and all peak shapes were good. The interference of serum impurities on the determination did not occur. The curves showed good linearity. The recoveries of 11 FFA were>75%,respectively. The relative standard deviations(RSD) of intra-day and inter-day were<10%,respectively. Conclusions　The method is sensitive,accurate,easy to operate and time-saving. It provides referential basis to the determination of serum FFA in coronary disease.

Cause analysis and countermeasure of the rejection of blood specimens

ZHU Jing, ZHAO Ying, WANG Beili, WU Jiong, SONG Binbin, ZHANG Chunyan, GUO Wei, PAN Baishen

2014, 29(3):  288-292.  DOI: 10.3969/j.issn.1673-8640.2014.03.021

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Objective　To analyze the cause and countermeasure of the rejection of blood specimens，and to find solutions to decrease the number of rejected blood specimens for laboratory quality improvement. Methods　A total of 5 933 rejected blood specimens were analyzed retrospectively from 2008 to 2012 in Department of Clinical Laboratory，Zhongshan Hospital，Fudan University. Results were shown by the frequency of rejected blood specimens in which Pearson χ² test was used to evaluate the percentages of clotted specimens and insufficient specimens with different types of tubes. Results　The frequencies of rejected blood specimens from 2008 to 2012 were 1.49 ‰，0.76 ‰，0.52 ‰，0.50‰ and 0.47‰，respectively，with decreasing tendency. The 6 major causes of rejection were clotted blood specimen，insufficient blood volume，inappropriate anticoagulant，error in sample identification，specimen transfer overtime and repeated blood collection. Clotted blood specimen and insufficient blood volume were common in sodium citrate anticoagulant tube specimens compared with others. Conclusions　Clinical laboratories should strengthen the ties with the nurses and clinical doctors whom would be informed with the rejection of blood samples in time，in order to figure out and carry out the solutions，reduce the number of rejected blood specimens and improve the pre-analytical quality.

Laboratory diagnosis for sepsis and its research advance

CHEN Hongwei, LIANG Dongyu, HOU Yanqiang.

2014, 29(3):  297-300.  DOI: 10.3969/j.issn.1673-8640.2014.03.024

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Sepsis,a common deadly systemic infection caused by a variety of pathogens,has some clinical symptoms similar to the systemic inflammatory response syndrome (SIRS). Currently,many parameters have been used for the clinical diagnosis of sepsis,however,none of them has gained unanimous acceptance as a specific parameter for sepsis. Despite of the development in diagnositic and therapeutic techniques,the incidence and mortality still remain high. This article summarizes widely applied or hot research parameter for the clinical diagnosis.