Comparison of double antigen sandwich and indirect enzyme-linked immunosorbent assay for the diagnosis of hepatitis C virus antibodies.

BACKGROUND: The aim of this study is to compare double-antigen sandwich enzyme-linked immunosorbent assay (ELISA) and indirect ELISA in the diagnosis of hepatitis C virus（HCV）infection. METHODS AND MATERIALS: A total of 176 samples from the Tumor Hospital Affiliated to Xin Jiang Medical University were utilized to comparison. All serum samples were tested using double-antigen sandwich ELISA and indirect ELISA. Cohen's kappa statistics were used to assess the agreement between the two assays, and multivariate analysis was used to evaluate risk factors for the discordance between the double-antigen ELISA and indirect ELISA. RESULTS: The positivities of indirect ELISA (Beijing Wantai), double-antigen sandwich ELISA (Beijing Wantai), and indirect ELISA (Beijing Jinhao) were 74.43%, 68.75%, and 73.30%, respectively. The agreement between the indirect ELISA (Beijing Wantai) and double-antigen sandwich ELISA (Beijing Wantai) was high (κ = 0.829;P < .001), and the agreement between the double-antigen sandwich ELISA (Beijing Wantai) and indirect ELISA (Beijing Jinhao) was high (κ = 0.847;P < .001). Variables associated with discordant results between the double-antigen sandwich and indirect ELISA in multivariate analysis were as follows: female (OR:1.462; P < .05), age (<35 years old; OR:3.667; P < .05), and cancer (suffer from malignant tumor; OR:3.621; P < .05). CONCLUSION: In detection of HCV, high agreement was found between the double-antigen sandwich ELISA and indirect ELISA. Female, younger age, and suffer from malignant tumor were significant risk factors for the discordance. Based on double-antigen sandwich ELISA has distinct methodological advantages over indirect ELISA. It is recommended for the diagnosis of HCV infection.

[Methylation Chip Screening and Verification of Differential Genes Related to Tuberculosis Infection].

OBJECTIVE: To screen the genes with significant changes in DNA methylation level in active tuberculosis patients, we used the methylation chips and expanded the sample size to verify candidate genes. METHODS: ① This study enrolled 9 cases of active tuberculosis patients, 3 cases of latent tuberculosis patients and 3 cases of healthy controls whose age and gender were all matched. Genome DNA was extracted from peripheral blood mononuclear cell in blood samples collected from these candidates, and bisulfite conversion treatment was then conducted. After hybridization with the Illumina HD 450K Infinium Mehtylation BeadChip, the results were compared between patients group and control group, and GO and KEGG pathway analyses were performed to evaluate the function of differentially expressed genes. ② We further enrolled 60 cases of active tuberculosis patients and 60 cases of health controls (age-and gender-matched), DNA was extracted from their peripheral blood and also followed bisulfite conversion treatment. Pyrosequencing method was used to detect the methylation levels of candidate genes (IFNGR2, PTPN6, CRK1, ATP6V0B, WIF1, DKK1 and SFRP1) screened by gene chip. RESULTS: Compared with healthy controls, the fragments in the patients that showed low methylation change accounted for the vast majority. Most of the methylation differential fragments (DMRs) were located in the main body region, followed by the upstream region of transcription initiation site, and the lowest DMRs distribution area was 3´UTR area. GO and Pathway analysis showed that the functions of the differentially methylated regions related genes are mainly enriched in the biological processes of the regulation of leukocyte differentiation, apoptosis, cytokine regulation and inflammatory response which are closely related to tuberculosis. There were 32 CpG sites involved in the verified 7 tuberculosis related genes, and 16 CpG locus showed significant difference (P<0.05), they were distributed in 6 genes: PTPN6, WIF1, CRK1, SFRP1, DKK1 and IFNGR2.Of these genes with significant difference, PTPN6 genes showed hypermethylation status and WIF1, CRK1, SFRP1, DKK1 and IFNGR2 genes exhibited demethylation status in the patients group compared to the health controls. SFRP1 and CRK-1 mRNA up-regulated in the patients group compared with health controls. CONCLUSION: In the course of MTB infection, the methylation status of genomic DNA is altered, and most of the differentially methylated regions (DMRs) are showed status of demethylation. The expressions ofSFRP1and CRK-1gene up-regulate in tuberculosis infection.

The Leukocyte VCS Parameters Compared with Procalcitonin, Interleukin-6, and Soluble Hemoglobin Scavenger Receptor sCD163 for Prediction of Sepsis in Patients with Cirrhosis.

BACKGROUND: Patients with liver cirrhosis have a high risk of sepsis and a poor prognosis. Recently, a new standard for sepsis (Sepsis-3) has been proposed in the general population. The Coulter Lh 750 hematology analyzer can evaluate mean volume, conductivity, scatter, and distribution width of leukocyte. We tried to use Sepsis-3 criteria to study the diagnostic value of volume, conductivity, and scattering (VCS) parameters in sepsis and infection in patients with liver cirrhosis compared with traditional infection markers (PCT, IL-6, sCDl63). METHODS: A blinded, cohort study was conducted in three different ED populations within three affiliated hospitals. A total of 249 patients with liver cirrhosis were enrolled in the study. According to the "Sepsis-3" consensus criteria, clinical history, and laboratory examination, the subjects were divided into sepsis (n = 54), patients with infections (n = 95), and patients without systemic infections (n = 100). The blood samples of the patients were collected at the time of ED admission and were evaluated for the detection of sepsis. RESULTS: The differences of MNV, MNS, MMV, MMS, MLV, NDW, and MDW in the three groups were statistically significant. In the diagnosis of sepsis in patients with liver cirrhosis, the sensitivity of combined detection of MMV and MDW was 88.89%; the specificity was 74%. This sensitivity was significantly better than the 83.3% achieved using 0.97 mg/L as the cutoff for sCD163. In the diagnosis of infection in cirrhosis, the sensitivity of combination of MNV and MMS was increased to 86.32%; the specificity was 92%. The sensitivity was the same as that achieved by using 0.31 ng/mL as the cutoff value of PCT, but the specificity increased. CONCLUSION: The leukocyte VCS parameter could be potential parameters for indicating sepsis and infection in patients with liver cirrhosis. The combined detection of MMV and MDW seemed to be helpful for the diagnosis of sepsis in these patients, and the combination of MNV and MMS could better indicate infection for them.

The relationship between the PD-1/PD-L1 pathway and DNA mismatch repair in cervical cancer and its clinical significance.

BACKGROUND: According to recent clinical observations, deficient DNA mismatch repair (dMMR) is capable of improving antitumor effects of the PD-1/PD-L1 pathway, suggesting that dMMR may act as a prognostic indicator of PD-1/PD-L1 antibody drugs. In this study, we examined the dMMR and PD-1/PD-L1 expression, as well as explored the correlation of dMMR status with PD-1/PD-L1 expression in cervical cancer patients, in order to optimize cervical cancer patient selection for PD-1/PD-L1 antibody drug treatment, which is helpful to avoid adverse effects and keep costs manageable. METHODS: Sixty-six tissue samples from patients with squamous cell carcinoma were collected, and data of their clinical characteristics were also gathered. Based on these samples, the expression levels of MLH1, MSH2, and PD-L1 in cancer cells were tested by immunohistochemical assay (IHC). Moreover, PD-1/PD-L1 expression in tumor-invading lymphocytes (TILs) was detected by IHC as well. Six single-nucleotide-repeat markers of microsatellite instability (MSI), including NR-27, MONO-27, BAT-25, NR-24, NR-21, and BAT-26, were tested by capillary electrophoresis sequencer analysis. According to expression of MLH1, MSH2 and the MSI test, all 66 cases were divided into dMMR or proficient DNA mismatch repair (pMMR) groups. The comparisons of dMMR and PD-L1 in cancer cells and of PD-1/PD-L1 in TILs were conducted categorized by age, childbearing history, history of abortion, ethnicity, and cancer cell differentiation subgroup. Furthermore, PD-L1 levels in cancer cells and PD-1/PD-L1 in TILs were analyzed and compared in both dMMR and pMMR subgroups. RESULTS: Of the patient samples, 25.8% were associated with dMMR. PD-L1 in cancer cells, PD-L1 in TILs, and PD-1 in TILs took up 59.1%, 47.0%, and 60.6%, respectively. The data indicated that both dMMR and PD-L1 overexpression resulted from lower cancer differentiation, more incidences of childbearing, and a history of abortion. Abortion could significantly increase PD-1 expression levels in TILs. Additionally, more incidence of childbearing or older age (35-55 years) was able to upregulate PD-L1 expression in TILs. Statistical difference of PD-L1 in cancer cells could be observed between dMMR and pMMR subgroups. In the dMMR group, PD-L1 in cancer cells and PD-1 in TILs had no correlation (rs=0.161, p=0.537), but in the pMMR group, they had good correlation (rs=0.645, p<0.001). CONCLUSION: According to prior studies and our own experiments, PD-L1 in both cancer cells and TILs and PD-1 in TILs are widely observed in cervical cancer patients, indicating that there may be potential to apply PD-1/PD-L1 antibody drugs in cervical cancer. dMMR patients are associated with higher PD-L1 expression compared with pMMR ones, which suggested that PD-1/PD-L1 antibody drugs may work well in dMMR cervical cancer patients. Moreover, in patients with more incidences of childbearing or abortion, dMMR may be a molecular detection target for clinical application of PD-1/PD-L1 antibody drugs.

[Bloodstream infections in southwestern China: 2012 Whire Union report on bacterial susceptibility to antibiotics].

OBJECTIVE: To test the susceptibility of blood culture isolates to antibiotics in a sample of hospitals in Southwestern China in 2012. METHODS: Blood culture samples were taken and tested from the patients under the Whire Union surveillance in three tertiary hospitals in Southwestern China in 2012. We performed antimicrobial susceptibility analyses on the blood culture isolates using WHONET5. 5 and SPSS19.0. RESULTS: A total of 1745 isolates were identified: 877 Gram-positive (50.26%); 868 Gram-negative (49.74%). Coagulase-negative Staphylococcius (CNS, 541 strains, 31.00%), Escherichia coli (ECO, 379 strains, 21.72%), Klebsiella spp. (170 strains, 9.74%), Enterococcus spp. (143 strains, 8.20%) and Staphylococcus aureus (SAU, 128 strains, 7.34%) comprised the majority of isolates. Methicillin-resistant was found in 37.5% of S. aureus and 85.4% of CNS, respectively. Zero and 6.3% resistance to vancomycin were found for Enterococcus faecalis and E. faecium, respectively,compared with a 1.4% and 7.9% resistance to linezolid, respectively. Imipenem-resistance occurred in 0.7% E. coli, 3.6% Klebsiella spp., and 11.1% Enterobacter spp.. Nonfermenters were highly resistant to carbapenems, with an imipenem resistance rate of 25.0% in P. aeruginosa and 79.1% in Acinetobacter baumannii. Antimicrobial usage and critical-care units were identified as risk factors for MRSA and multi-drug-resistant A. baumannii infections. Patients undergoing tracheotomy/endotracheal intubation were likely to develop multi-drug-resistant A. baumannii infections than others. CONCLUSION: CNS, E. coli,Klebsiella spp., Enterococcus spp. and S. aureus were the predominant organisms in bloodstream infections in Southwestern China. Low methicillin-resistant rate was found in Staphylococcus aureus. But vancomycin-resistant rate was high in E. faecium. Imipenem-resistant rates varied in Enterobacteriaceae: higher in non-fermenting bacteria especially in Acinetobacter spp. Use of abtimicrobial drugs and invasive procedures can lead to development of antimicrobial drugs-resistance.

DNA ploidy of cervical epithelial cells should be a cure criterion of high-risk HPV infection in Xinjiang Uygur women.

BACKGROUND: The Uygur women have the highest incidence of cervical cancer in all Chinese ethnic groups. The research was conducted to explore whether DNA ploidy could be the prognostic indicator of human papillomavirus (HPV) infection in Xinjiang Uygur women. METHODS: Case data and cervical exfoliated cell samples from 326 Uygur women. The DNA ploidy was analyzed by flow cytometry. The flow-through hybridization and gene chip (FHGC) for HPV type test then divided the cases into negative HPV group, non high-risk HPV infection group, single high-risk HPV infection group, and multiple high-risk HPV infection group. Lastly, 113 cases from 273 HPV infection cases were followed up. RESULTS: The 16-type HPV had the highest rate in all genotypes infection; 16/18-type HPV mixed infection was the most common type in multiple high-risk HPV infection group. Abnormal DNA ploidy happened along with the seriousness of HPV infection. Compared with the HPV negative group, DNA heteroploid appeared 12.750 times and 22.705 times, respectively, in single high-risk HPV and multiple high-risk HPV infection groups. Followed up 1 year later, the DNA index, S-phase cells' peak percentage and heteroploid of cervical exfoliated cells significantly reduced in single and multiple high-risk HPV infection patients, but in nine patients negative HPV infection and DNA heteroploid still existed. CONCLUSION: The finally cure criterion of high-risk HPV infection should include the negative HPV test and normal DNA ploidy analysis. It was useful to prevent and cure cervical lesions in Xinjiang Uygur women through high-risk HPV test and DNA ploidy analysis. The transient infection and persistent infection in Xinjiang Uygur women should be taken as further research.

[Separation and purification and structural characterization of baicalein].

The root of scutellaria baicalensis georgi that contains a variety of flavonoids is a very old and well-known drug in traditional Chinese medicine, which is widely used for treatment of bronchitis, tumors and inflammatory diseases. The baicalein is the main active component from traditional Chinese medicine-scutellaria baicalensis georgi. It is a very significance research work that the baicalein was separated and purified, and its composition and molecular structure are analyzed and determined for the pharmacology study of Chinese medicine-scutellaria baicalensis georgi. The main works in this paper are as follows. Powdered roots (100 g) were extracted with methanol by three times, each time for 48 hours. The crude extracts were purified by polyamide column chromatography and CH3Cl-C2H5OH gradient desorption. A short yellow prismatic crystal was acquired by recrystallizing technique and its composition and molecular structure were characterized by color reactions and spectral analysis methods as FTIR, UV-Vis, MS and 1H NMR, 13C-NMR. The FTIR spectrum appears the absorption bands for hydroxyls, pyrone carbonyl, aromatic C=C bond and singly substituted phenyl. The characteristic absorption peaks and the vibration modes in FTIR spectrum were identified as corresponding groups. The UV-Vis spectrum in methanol solution and the mix solution of methanol with 5 diagnostic reagents, NaOMe, NaOAc, NaOAc/H3BO3, AlCl3, AlCl3/HCl, respectively indicate that the yellow prismatic crystal is flavone with 5-hydroxyl, 4-carbonyl and 5,6,7- or 5,7,8-trihydroxyls on ring A. The structure of the crystal was characterized by three different MS. The results of FAB-, ESI- and EI-MS show that it is not a flavone glocuside but the flavone with three phenyl hydroxyls on ring A, and no OH group and other substituted groups on ring B. The molecular ion and fragment ions are identified by MS, which include such as m/z 270 M+, m/z 242 [M-CO]+, m/z 168 A, m/z 140 [A1-CO]+, m/z 105 B, m/z 102 B, m/z 77 [B2-CO]+, respectively. 13C-NMR (DMSO-d6) exhibits the signals of the fifteen carbon atoms, nine oxygenous aromatic C, five non-oxygenous aromatic C and a carbonyl C. 1H-NMR(DMSO-d6 + D2O, DMSO-d6 )indicates the presence of C-5, C-6, C-7 hydroxyl protons, which is consistent with the results of UV spectrum. The signals for C-2',6' hydroxyls appear at delta = 8.055 as a doublet peak with spin-spin coupling constant 6.0 Hz. The other signals were ascribed to the corresponding H or C atoms in the compound. The results of FTIR, UV-Vis, MS, 1H NMR, 13C-NMR spectroscopy characterization show that crystal is the 5,6,7-trihydroxy-flavone, that is baicalein, and the molecular formula is C15H10O5.

[UV-Vis spectroscopic characterization of active components from Scutellaria baicalensis georgi].

The molecular structures of three active components from scutellaria baicalensis have been studied by ultraviolet-visible spectra. The index flavonoid structures and substituted positions were deduced by analyzing the UV-Vis spectra in the methanol solution of three active components and the methanol solution with 5 diagnostic reagents, NaOMe, NaOAc, NaOAc/H3BO3, AlCl3, AlCl3/HCl respectively, which provided strong evidences for the structural characterization of the active components from natural products.