Objective To evaluate the effect of immune fluorescence chromatography on glycosylated hemoglobin (HbA1c). Methods The precision of immune fluorescence chromatography was evaluated with samples of 6.0% and 8.0% fixed value. Group of High performance liquid chromatography (HPLC) as control, HbA1c for 200 samples of EDTA-K2 anti-coagulated whole blood were detected by immune fluorescence chromatography to synchronous blinded trial. Results As to the precise of immune fluorescence chromatography in the samples of 6% and 8%, values of coefficient of variation were 5.1% and 5.3%, respectively. The linear regression equation of immune fluorescence chromatography and HPLC was Y=-0.110+1.021X and the correlation coefficient was 0.982. 6.0% and 8.0% as the cut-off value, kappa values were 0.950 (P < 0.001) and 0.922 (P < 0.001), respectively. Conclusion Immune fluorescence chromatography and HPLC is consistent with detection of HbA1c, which can be used for clinical detection of HbA1c.

Objective To study the expressions of osteopontin (OPN),caspase-3 and mt-P53 proteins, and their relationship in gliomas. Methods Seventy gliomas specimens of patients (glioma group) were selected, and 10 samples of non-glioma brain tissue were used as control group. The SP method was used to detect the positive rates of protein expressions of OPN, caspase-3 and mt-P53 between two groups. The relationship between protein expressions of OPN, caspase-3 and mt-P53 in gliomas and grade of gliomas were detected by Western blot assay. Spearman rank correlation was compared between the positive expression of OPN, caspase-3 and rate mt-P53. Results The positive expression rates of OPN and mt-P53 were significantly higher in glioma group (64.29%and 60%) than those of control group (no positive expression), but the positive expression rate of caspase-3 was significantly lower than that of control group (47.14%vs. 90%, P<0.05). There were no significant differences in OPN, caspase-3 and mt-P53 expressions between different gender, age, tumor size and tumor position (P>0.05). The higher the WHO classification, the higher the positive expression rates of OPN and mt-P53 (P<0.001), and the lower the positive expression rate of caspase-3 (P<0.001). With the increased level of glioma grade, OPN and mt-P53 protein levels were increased, but caspase-3 protein expression level was decreased. There was a negatively correlation between OPN and the positive expression of caspase-3, but there was a positive correlation between OPN and the expression of mt-P53 (rs=-0.720 and 0.722, P<0.05). There was a negative correlation between caspase-3 and mt-P53 expressions (rs=-0.556, P<0.05). Conclusion The higher the WHO classification, the higher the positive expression rates of OPN and mt-P53, while the lower the positive expression rate of caspase-3. The study reveals that OPN, caspase-3 and mt-P53 expressions are associated with the occurrence and the progress of gliomas. The combined detection of them can contribute to the judgment of biological behavior of gliomas.

Objective To establish an assay for the detection of Streptococcus pneumoniae by real-time fluorescence quantititive polymerase chain reaction (PCR).Methods Special primers and probe for the autolysin A (lytA)gene were designed.The sensitivity and specificity of primers and probe were studied,and cut-off of cycle threshold was assayed.158 clinical specimens were confirmed by real-time fluorescence quantitative PCR and bacterial culture method.Results Primer and probe design for LytA gene could sensitively detect serotype Streptococcus pneumoniae strains of common pathogenic,and the sensitivity was 100 copies.Among 35 strains of Streptococcus pneumoniae,34 cases were detected to be positive for Streptococcus pneumoniae by real-time fluorescence quantitative PCR,while 1 case was detected to be negative;among 15 strains of non-Streptococcus pneumoniae, all were detected to be negative.Among the 158 clinical sputum specimens,34 cases with Streptococcus pneumoniae were detected by real-time fluorescence quantitative PCR,while only 10 cases with Streptococcus pneumoniae were detected by the culture method.White blood cells count and time in hospital of cases with Streptococcus pneumoniae were higher than those of cases without Streptococcus pneumoniae (P <0.05 ). Conclusion Real-time fluorescence quantitative PCR is a sensitive and specific assay for the detection of Streptococcus pneumoniae.It can be used for the diagnosis of Streptococcus pneumoniae.

Objective To develop a rapid quantitative detecting assay for point-of-care testing ( POCT ) of N-terminal pro-brain natriuretic peptide ( NT-proBNP ) in serum by the fluorescence immunochromatographic technology.Methods Applying double-antibody sandwich assay to establish the quantitative NT-proBNP kit.The performance of quantitative NT-proBNP kit was evaluated by the sensitivity , specificity, accuracy, precision, stability and clinical effectiveness.It compared the research kit and conference kit by the parallel experience in the 1 056(605 males, 451 females)serum specimen collected from Guangdong Provincial People′s Hospital, Sun Yat-sen Memorial Hospital and Children′s Hospital of Zhengzhou between February 2013 to April 2014.Statistical significance of the results was assessed by correlation analysis , linear regression , receive operating characteristic ( ROC) curve analysis , negative and positive consistent.Results The report range of the NT-proBNP kit was 18-35 000 ng/L.The coefficient of variation ( CV) values for low , median and high concentration calibrators respectively were all less than 15%.Common interfering substances in human serum specimens such as bilirubin , triglyceride and cholesterol were found no significant affect on NT-proBNP antigen detection and the CV were no more than 15%.According to the results of detection for calibrators , the shelf time of the NT-proBNP diagnostic kit should be longer than 12 months.The NT-proBNP kit and reference kit had good correlation ( Y=1.048 9X developed reference +121.54, R2 =0.956 6, n=1 056) to detect the target protein through the parallel experiments and the deviation of the quantitative results of clinical serum samples showed no statistical significance (Z=0.88, P=0.379>0.05).The clinical assays of two different diagnostic kits showed good consistency based on the ROC curve evaluation which is compared by two cut-off values (300 and 450 ng/L).The areas under ROC curve were 0.981 and 0.978 respectively.Conclusions A novel NT-proBNP chromatographic quantitative immunofluorescence detection method was developed in this study .The performance evaluation data indicated that the kit is suitable for rapid detection of serum NT -proBNP.

Objective To investigate the correlation of the 1896 and 1899 mutations of hepatitis B virus (HBV)with the conversion of e antigen in serum and the progression of the disease. Methods 238 serum samples from the patients with HBsAg positive for over six months and HBV-DNA copy number > 5.0 × 102 IU/mL were collected,and the sequence analysis was used to analyze the nucleotide sequences of the 1896 and 1899 sites in the pre-C region of HBV. At the same time,the relevant clinical data and the expressions of HBeAg were collected,followed by Spearman correlation analysis and chi square test with SPSS 20.0. Results Both 1896 and 1899 sites in the pre-C region of HBV were mutated,and the base G was A,which was closely related to the expression of e antigen(P<0.05). Both G1896A and G1899A promoted the e antigen serological conversion ,and the e antigen serological conversion of G1899A was higher than that of G1896A. G1899A was associated with HBV related disease progression (correlation coefficient 0.280,P < 0.05),especially with the incurrence of HCC. Conclusions G1896A and G1899A in the pre-C region of HBV can promote the serological conversion of e antigen.

OBJECTIVETo observe the effect of mesenchymal stem cells derived from umbilical cord (UC-MSCs) on natural killer (NK) cells-mediated cytotoxicity against dendritic cells (DCs) and explore the mechanism.

METHODSMSCs were isolated from human umbilical cord by collagen digestion and cultured in vitro. NK cells were separated from healthy human peripheral blood by magnetic bead sorting. Mononuclear cells from healthy human peripheral blood were cultured in the presence of granulocyte and macrophage colony stimulating factor (GM-CSF) and interleukin-4 (IL-4) to obtain the immature DCs. The DCs were then co-cultured with UC-MSCs in the presence of tumor necrosis factor α (TNFα) for 2 days, and the expressions of CD11c and CD86 on DCs and IL-12 level in the culture medium was detected using flow cytometry and ELISA, respectively. The cytotoxicity of NK cells against DCs was analyzed by LDH-releasing assay, and the expressions of ligands for killer activator receptor (MICA/B and ULBP1-3) on the DCs were detected with flow cytometry.

RESULTSCompared with the cytokine-induced DCs, the DCs induced by co-culture with UC-MSCs showed an identical CD11c expression but lowered CD86 expression and IL-12 secretion. The natural killer cells produced a stronger cytotoxicity against UC-MSCs-induced DCs than against cytokine-induced DCs. The UC-MSCs-induced DCs also showed increased expressions of MICA and MICB on the surface.

CONCLUSIONUC-MSCs can enhance NK cells-mediated cytotoxicity against DCs possibly by inhibiting DC maturation and up-regulating the ligands for killer activator receptor on the surface of the DCs.

Cells, Cultured ; Cytotoxicity, Immunologic ; immunology ; Dendritic Cells ; cytology ; immunology ; Humans ; Killer Cells, Natural ; cytology ; Mesenchymal Stromal Cells ; cytology ; Umbilical Cord ; cytology

Objective To screen and identify B cell epitopes in SAG1, SAG2, SAG3, GRA1, GRA6 and P35 antigens of Toxoplasma gondii. Methods The indexes such as hydrophilicity, accessibility, flexibility, secondary structure and polarity of the 6 antigen moleculars above mentioned were analyzed by BioSun system. Two B cell epitopes with high antigenicity from each antigen molecular were selected, and the total twelve pairs of oligonucleotide chains were designed according to the 12 B cell epitopes’ sequence and synthesized, then cloned into plasmid pET-32c. The 12 fragment B cell epitopes were expressed and the expressed fusion proteins were identified with Western blot. Results Twelve B cell epitopes from 6 Toxoplasma antigens (two from each antigen) were predicted and selected. The epitope genes were successfully cloned into pET-32c and expressed. Western blot results showed that 3 of 12 expressed fusion proteins could be recognized by the immunized rabbit sera with soluble antigen of Toxoplasma gondii, but not by the unimmunized rabbit sera Conclusion Three B cell epitopes of Toxoplasma[with potential diagnostic value are obtained.

Objective To investigate the effect of prevascularized tissue-engineered bone graft on regeneration of femoral bone defects in rats.Methods Models of femoral bone defect were created at the bilateral hind limbs of 20 healthy female 10 week-old rats which were divided into 2 even groups randomly (n =10).In group A,conventional tissue-engineered bone grafts were transplanted into the femoral bone defects;in group B,tissue-engineered bone grafts and vascular bundles were implanted into the femoral defects.At 1,4 and 8 weeks after operation,3 rats were sacrificed each time in each group to harvest samples.The remaining one in each group served as a spare animal.Regeneration of bone defects and degradation of scaffolds were assessed by radiologic modality and hematein eosin staining.Results At week 1,the new bone ratio (BV/TV) was 5.47％ ± 1.90％ in group A and 8.49％ ± 1.26％ in group B,showing no significant difference (P ＞ 0.05);at weeks 4 & 8,the BV/TV were 17.54％ ±2.04％ and 39.73％ ± 4.01％ in group A,significantly lower than those in group B (25.32％ ± 2.15％ and 53.22％ ± 2.94％) (P ＜ 0.05).At weeks 1 & 4,the scaffold degradation ratios (RSV/SV) were 97.33％ ± 2.52％ and 80.60％ ±4.00％,showing no significant differences from those in group B (95.67％ ±3.51％ and 75.22％ ±6.20％) (P ＞ 0.05).At week 8,the scaffold degradation ratio in group A (65.46％ ±4.51％) was significantly higher than that in group B (50.19％ ±4.91％) (P ＜ 0.05).At week 8,hematein eosin staining showed better integration of scaffolds with the femur,faster degradation of the interior scaffolds and greater osteogenetic activity in group B.Conclusion Prevascularization of tissue-engineered bone graft may increase new bone volume and scaffold degradation rate,promoting repair of femoral bone defects in rats.

Objective:To discuss the surgical approach and the clinical effect of laparoscopic enucleation for hepatic hemangioma(HH).Methods:Forty HH patients admitted in the Third Affiliated Hospital of Soochow University between Aug 2018 and Dec 2019 were analyzed. The technical knowhow involved is to make a good explosure of the pseudocapsular HH from underneath. Herewith a process of HH enucleatoin started.Results:All the 40 patients undergone total laparoscopic hepatectomy successfully.The operative time was (90.3±32.3)min, the intraoperative blood loss was (50±500)ml, the time of hepatic block was (12.5±35.4) min. The volume of postoperative drainage was 10-150 ml on the first postoperative day, and was gradually reduced to <30 ml on the third day after the operation. All patients were up and about from post-op day 2. The length of stay after operation was (5.5±2.7)d. There were significant differences in ALT、AST、TBIL and prealbumin in 1 and 3 days after surgery (seperately, t=-5.481, -4.182, -2.235, 9.722, all P<0.05), before back to normal on day 7 (seperately t=0.167, -1.392, 1.000, -2.531, all P>0.05). Liver function recovered to normal in 7 days after surgery. Conclusion:New approach breaks the traditional stripping method and makes the procedure simple, safe, lessly disturb the liver function.

Objective To systematically evaluate the biomechanical recovery of drilled holes in the femur in SD rats.Methods Eighteen female SD rats were randomized into 3 even groups (n =6).Models of 2-mm drilled holes in bilateral femurs were established in groups A and B with 2 holes on each side while no drilling was performed in group C.Samples were harvested in group A at postoperative 4 weeks,in group B at postoperative 8 weeks while at both 4 and 8 weeks in group C.The samples were evaluated in terms of linear elasticity (compression test),viscoelasticity (relaxation and creep tests) and durability (fatigue failure test).Micro-CT scan was performed to measure the bone volume fraction (BV/TV) and bone mineral density (BMD) of new bone.Sirus red staining was performed to measure regeneration of type Ⅰ collagen of new bone.Results The elasticity modulus,maximum load,compression strength and conditional yield limit in groups A were significantly lower than those in group B which were also significantly lower than those in group C (P ＜ 0.05).At 7,200 s,the relaxation (14.56 ±0.69 MPa) and creep variation (11.37％ ± 0.70％) in group A were significantly higher than those in group B (11.06 0.63 MPa and 8.98％ ± 0.40％) which were also significantly higher than those in group C (6.99 ±0.56 MPa and 5.10％ ±0.23％) (P ＜ 0.05).At the constant amplitude loads from 20 N to 200 N,from 20 N to 300 N and from 20 N to 400 N,the recycling numbers in group A (6,044.3 ±879.7,4,093.3 ±628.5 and 1,919.3 ±847.5) were significantly lower than those in group B (10,192.3 ± 1,109.1,6,750.6 ± 818.0 and 3,376.6 ± 671.3) which were also significantly lower than those in group C (28,068.3 ±2,702.6,11,788.3 ± 1,141.6 and 5,296.3 ± 735.0) (P ＜ 0.05).By micro-CT scan,the BVT and BMD in group A were significantly lower than those in group B which were also significantly lower than those in group C (P ＜ 0.05).The sirus red staining showed the type Ⅰ collagen in the bone defect area was completely regenerated in group B.Conclusion Systematic biomechanical measurements may actually detect the characteristics of biomechanical recovery of bone holes in SD rats,enriching the basic research on the bone damage repairing progress.