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Summary Objectives The definitive diagnosis of endobronchial tuberculosis (EBTB) is challenging because the disease manifests in various non-specific ways, and acid-fast bacilli (AFB) are often undetectable by sputum smear. The objective of this study was to evaluate the efficacy of real-time PCR of bronchoscopic biopsy specimens for the diagnosis of EBTB. Methods Real-time PCR amplification of Mycobacterium tuberculosis DNA in biopsy tissue from EBTB patients was performed prospectively. Diagnostic yields were compared for real-time PCR and for auramine O-stained sputum smears and bronchial brush smears. Whether diagnostic yield depended on bronchoscopic subtype of EBTB was also evaluated. Results Diagnostic yields were 4.1% (3/74) for sputum smear, 39.2% (29/74) for bronchial brush smear, and 89.2% (66/74) for real-time PCR. Real-time PCR melting curve analysis showed significantly higher yields than did AFB staining of bronchial brush smears for granular and caseating EBTB ( p Conclusions Real-time PCR detection of M. tuberculosis DNA in EBTB biopsy tissue is more sensitive than sputum smear and bronchial brush smear, including at early disease stages. This PCR method may be a useful adjunct to culture- and smear-based techniques to allow more rapid EBTB diagnosis and timelier treatment.

Background: Optimizing basic techniques in diagnostic bronchoscopy is important for improving medical services in developing countries. In this study, the optimal sequence of bronchial brushing relative to bronchial biopsy for lung cancer diagnosis was evaluated. Methods: A total of 420 patients with visible endobronchial tumors were prospectively and randomly enrolled in two groups: a pre-biopsy brushing group, receiving two brushings before biopsy; two brushings which performed afterwards; were set as self-control and compared with the pre-biopsy brushings as the intra-group comparison; and a post-biopsy brushing group, only receiving two brushings after biopsy, which were compared with the pre-biopsy brushings as the inter-group comparison. Diagnostic yield of brushing was compared before and after biopsy, and as well as for different tumor pathologies and bronchoscopic morphologies. The occurrence of treated bleeding which defined as bleeding needed further intervention with argon plasma coagulation and/or anti-coagulation drugs in two groups was also compared. Results: Only patients with a definitive cytological or histological diagnosis of lung cancer based on bronchoscopy or other confirmatory techniques were included. Patients were excluded if they had submucosal lesions, extrinsic compressions, pulmonary metastasis of extrapulmonary malignancies or uncommon non-small cell lung carcinoma (NSCLC). A total of 362 patients who met the inclusion criteria were analyzed. Diagnostic yield for pre-biopsy brushing (49.2%, 88/179) was significantly higher than for post-biopsy brushing within the same pre-biopsy brushing group (31.8%, 57/179) (P=0.007) as the intra-group comparison, and significantly higher than for post-biopsy brushing in the post group (30.6%, 56/183) (P

Background and objective: Transbronchial needle aspiration (TBNA) is a well-established diagnostic method that is underutilized due to the relatively high percentage of non-diagnostic samples and low success rates. This study was designed to evaluate the impact of liquid-based cytology test (LBC) on the diagnostic yield from TBNA. Methods: Ninety-seven consecutive patients who underwent TBNA due to significant mediastinal adenopathy were enrolled in the study. Each target site was aspirated four times, with the first and third aspirates being prepared for LBC and the second and fourth aspirates being reserved for conventional pick-and-smear (CPS) cytology. Results: Paired aspirates were obtained from 114 target sites, giving a total of 228 test samples from 97 consecutive patients. The overall diagnostic sensitivity of TBNA was 63.6% (56/88). The yields from small cell lung cancers were better than those from non-small cell lung cancers (P 20 mm on computed tomography were also associated with better yields than nodes with diameters of 10–20 mm (P = 0.001). The diagnostic sensitivity of TBNA was similar for each processing method—59.8% (61/102) for LBC and 64.7% (65/102) for CPS. Conclusions: LBC was not inferior to CPS with respect to diagnostic yields from TBNA, and enabled efficient pathological evaluation.

Inulinase is an enzyme that belongs to glycoside hydrolase family 32. It converts inulin into high-fructose syrups and fructoligosaccharides, both of which are widely used in pharmaceutical and food industries. In this study, the kcINU1 gene (GenBank accession number AF178979) encoding an exoinulinase was cloned from Kluyveromyces cicerisporus CBS4857 and expressed in Pichia pastoris X-33, yielding a maximum of 45.2 ± 0.6 U mL(-1) of inulinase activity of culture supernatant. The expressed inulinase was purified and characterized. The enzyme had an optimum temperature of 55 °C and an optimum pH of 4.5. It had a K m of 0.322 mM and a V max of 4317 μM min(-1) mg(-1) protein when inulin was used as a substrate. It retained nearly 90 % of the maximal activity after pre-incubation at 50 °C for 1 h or at pH ranging from 3.0 to 6.0 at 4 °C for 24 h, demonstrating that KcINU1 was stable at high temperature and low pH. Moreover, we constructed two KcINU1 mutants, Asp30Ala and Glu215Ala, by site-directed mutagenesis and confirmed via zymogram analysis that Asp-30 and Glu-215 of the enzyme were the catalytic active center. The present study has provided important information for understanding the catalytic mechanism of exoinulinase.

Many studies evaluated the correlations of CD133 expression with the clinical outcomes in patients treated with chemoradiotherapy (CRT) but yielded controversial results. This meta-analysis was performed to identify the impacts of CD133 expression on the prognosis of cancer patients treated with CRT. Electronic databases updated up to March 2014 were searched to find relevant studies. Relevant literatures without any language restrictions were searched via electronic databases as follows: Web of Science (1945 ~ 2013), the Cochrane Library Database (Issue 12, 2013), PubMed (1966 ~ 2013), EMBASE (1980 ~ 2013), CINAHL (1982 ~ 2013), and the Chinese Biomedical Database (CBM) (1982 ~ 2013). STATA software was used for the current meta-analysis. Hazard ratios (HR) and its corresponding 95% confidence interval (95% CI) were calculated. Six studies were identified with a total of 470 cancer patients treated with CRT. The meta-analysis results showed that CD133-positive patients had poorer overall survival (OS) than that of CD133-negative patients (HR = 2.13, 95% CI = 1.20 ~ 3.07, P 0.001). Furthermore, CD133-positive patients displayed shorter disease-free survival (DFS) than that of CD133-negative patients (HR = 1.74, 95% CI = 0.08 ~ 3.40, P = 0.039). Ethnicity-stratified analysis indicated that CD133 expression positively correlated with shorter OS among the Japanese, Chinese, and Spanish populations (all P 0.05). In conclusion, our findings suggest that CD133 expression may be positively correlated with poorer prognosis in cancer patients treated with CRT.

To explore the distribution of Chinese medicine (CM) syndrome types in primary liver cancer (PLC) and their differences of the survival time.From May 2007 to March 2009, recruited were 151 PLC inpatients at Department of Tumor, Guangdong Provincial Hospital of Traditional Chinese Medicine. Their survival time were statistically calculated. Patients' average survival time and median survival time were calculated using Kaplan-Meier method. The Log-rank test was used to analyze their differences of survival time among different CM syndrome types.The proportion of CM syndrome types in PLC patients were ranked from high to low as follows: mutual accumulation of dampness and blood stasis syndrome [MADBSS, 43.0% (65/151)], Gan-stagnation Pi-deficiency syndrome [GSPDS, 34.4% (52/151)], qi stagnation blood stasis syndrome [QSBSS, 9.3% (14/151)], retention of damp-heat syndrome [RDHS, 8.6%(13/151)], and Gan-Shen yin deficiency syndrome [GSYDS, 4.6% (7/ 151)]. The median survival time of different CM syndrome types were ranked from longer to shorter as follows: GSPDS (14.77 months), QSBSS (6.13 months), RDHS (5.27 months), MADBSS (4.78 months), and GSYDS (0.80 months). The mean survival times were ranked from longer to shorter as follows: GSPDS (12.40 months), QSBSS (8.84 months), MADBSS (6.99 months), RDHS (7.08 months), and GSYDS (0.72 months). There was statistical difference in the difference of the survival time among different CM syndrome types (P0.05).GSPDS and MADBSS were the most common CM syndrome types in PLC patients. There was difference in the survival time between GSPDS and MADBSS/between RDHS and GSYDS. There was difference in the survival time between MADBSS and GSYDS. Patients of GSPDS might get the best prognosis, while patients of GSYDS might get the poorest prognosis.

Transbronchial needle aspiration (TBNA) is a well-established diagnostic method that is underutilized due to the relatively high percentage of non-diagnostic samples and low success rates. This study was designed to evaluate the impact of liquid-based cytology test (LBC) on the diagnostic yield from TBNA.Ninety-seven consecutive patients who underwent TBNA due to significant mediastinal adenopathy were enrolled in the study. Each target site was aspirated four times, with the first and third aspirates being prepared for LBC and the second and fourth aspirates being reserved for conventional pick-and-smear (CPS) cytology.Paired aspirates were obtained from 114 target sites, giving a total of 228 test samples from 97 consecutive patients. The overall diagnostic sensitivity of TBNA was 63.6% (56/88). The yields from small cell lung cancers were better than those from non-small cell lung cancers (P0.05), and TBNA of subcarinal nodes provided better diagnostic yields (P0.05). Nodal diameters20 mm on computed tomography were also associated with better yields than nodes with diameters of 10-20 mm (P = 0.001). The diagnostic sensitivity of TBNA was similar for each processing method-59.8% (61/102) for LBC and 64.7% (65/102) for CPS.LBC was not inferior to CPS with respect to diagnostic yields from TBNA, and enabled efficient pathological evaluation.

Many studies evaluated the correlations of CD133 expression with the clinical outcomes in patients treated with chemoradiotherapy (CRT) but yielded controversial results. This meta-analysis was performed to identify the impacts of CD133 expression on the prognosis of cancer patients treated with CRT. Electronic databases updated up to March 2014 were searched to find relevant studies. Relevant literatures without any language restrictions were searched via electronic databases as follows: Web of Science (1945 ~ 2013), the Cochrane Library Database (Issue 12, 2013), PubMed (1966 ~ 2013), EMBASE (1980 ~ 2013), CINAHL (1982 ~ 2013), and the Chinese Biomedical Database (CBM) (1982 ~ 2013). STATA software was used for the current meta-analysis. Hazard ratios (HR) and its corresponding 95 % confidence interval (95 % CI) were calculated. Six studies were identified with a total of 470 cancer patients treated with CRT. The meta-analysis results showed that CD133-positive patients had poorer overall survival (OS) than that of CD133-negative patients (HR = 2.13, 95 % CI = 1.20 ~ 3.07, P

A new method for construction of a cloning vector (T-vector) for direct ligation with PCR products was described. The T-vector derived from pUC118 in which the unique restriction site of Eam1105 I in the region of Amp(r) gene was deleted and an artificial DNA fragment flanking two Eam1105 I was introduced at the site of BamHI. The modified vector was named as pUC118E. A T-vector with 3' over hang end of a single T can be obtained via digesting of pUC118E with Eam1105I. PCR products can be easily cloned with this T-vector.

To investigate the clinical effect of the comprehensive treatment to acute stage of attack.On the basis of the previous observation, the study of the randomization control with general treatment, treatment on acupuncture and western medical treatment were carried out. A comprehensive treatment on overall traditional Chinese medical differentiation according to the superiority of every treatment was assessed and evaluated in 522 patients with attack.The comprehensive treatment of cerebral infarction was superior to the western medicine treatment. General treatment, treatment on traditional Chinese medical differentiation, acupuncture group revealed different improvement on neural function, daily viability, cognitive function in various extent.The comprehensive treatment that based on overall traditional Chinese medical differentiation has advantage and characteristic. It has positive combined action to the attack and relevant to clinical setting, easier to popularization and application. Various appraising amount form has different evaluating effects in different stage.

This paper describes a novel method by using hierarchical structure neural network for unusual sounds recognition. Above all, it discusses the principle of the inspecting system against theft that is used in pipeline transportation. Then the paper presents the structure of neural network, the sampling and the neural network train. The system achieves inspection preventing theft and caution by sound detecting, processing and recognizing. It is attested through actual application that the system capability is stable, the correct recognition ratio attains to 100%, and the error warning ratio is below 1%. In addition, the system can be expanded according to requirement.

A single-chain antibody (ScFv) phage display library was created by cloning antigen-binding regions of V(H) (variable domain) and V(L) gene repertoires as fusion proteins with a minor coat protein of filamentous phage, from which high affinity completely humanized ScFv against PreS1 of hepatitis B virus could be screened and characterized.A combinatorial library of phage-display human ScFv genes, which were derived from peripheral blood lymphocytes immunized by peptide PreS1 in vitro, was constructed. The library contained 7 x 10(8) clones.After 3 rounds panning, a high affinity (K=10(-7)-10(-8)mol/L) ScFv specific to PreS1 was obtained. Sequence analysis showed that the V(H) belonged to the V(H)4 family and V(lambda) to V(lambda)4.The described ScFv may provide a more satisfactory therapy. This application further illustrates that the method of in vitro antigen stimulation is expeditious for the source of human immune antibody library.

Screening and characterizing high affinity completely humanized single-chain antibody (ScFv) against PreS1 of hepatitis B virus.A combinatorial library of phage-displayed human ScFv, genes of which were derived from peripheral blood lymphocytes immunized by PreS1 of Hepatitis B Virus in vitro, was constructed. The library contained 7 10(8) clones.After 3 rounds panning, a high affinity (K=10(7) to 10(8) mol/L) ScFv specific to PreS1 was obtained. The V(H) belonged to human V(H4) family, and V(1) to V(4) by sequence analysis.This study suggests that the method of antigen stimulation in vitro is an expeditious way for the source of human immune antibody. And the ScFv may provide a more satisfactory therapy.

Total RNAs were extracted from the venom gland of the snake Gloydius ussuriensis. The cDNA of gussurobin, a thrombin-like enzyme from Gloydius ussuriensis, was cloned and amplified by RT-PCR. Assay of the nucleotide sequence of the cDNA allowed postulation of the complete amino acid sequence for Gloydius ussuriensis, Chinese Viperdae. Its amino acid sequence exhibits significant homology with that of other snake thrombin-like enzymes. The cDNA of gussurobin was inserted into the vector pPIC 9K and expressed successfully in Pichia pastoris, strain GS115. The recombinant gussurobin was separated and purified from 500 ml culture and showed one band on SDS-PAGE.