Objective To develop a double antibody sandwich ELISA for the determination of the Coxsackievirus A6(CVA6) antigen content, and to verify and preliminarily apply it for the CV-A6 production process and the quality control of tetravalent hand-foot-mouth disease(HFMD) vaccine.Methods Paired antibodies were screened and double antibody sandwich ELISA was developed by using purified CV-A6 sheep polyclonal antibody as coating antibody and mouse anti-CVA6 monoclonal antibody as detection antibody. The experimental conditions were optimized, and the linearity, range,specificity, accuracy, precision and robustness of the method were verified. The developed method was used to detect the antigen content of CV-A6 antigen production process samples and final products.Results Both of the sheep polyclonal antibody and monoclonal antibody 16D1-1E5 had neutralizing antibody activity, while the other three monoclonal antibodies showed no neutralizing antibody activity. The ELISA titers of all antibodies were not lower than 1. 00 × 105. The monoclonal antibody 2B6-4F9 was IgG3 subtype, while the other three monoclonal antibodies were IgG2a subtype. Using sheep polyclonal antibody as coating antibody and monoclonal antibody 16D1-1E5 as detection antibody for pairing, the method was developed.The dilution ratios of coating antibody and detection antibody were 1∶2 000-1∶6 000 and 1∶4 000-1∶6 000 respectively.The linear range of the developed method was 0. 664-21. 263 U/mL, with the R2≥ 0. 99. The method could only detect CVA6 antigen specifically and had no cross-reaction with CV-A10, CV-A16 and enterovirus 71(EV-71) antigens. M199,DMEM, Vero cell protein and bovine serum protein probably existing in the production process had no interference with this method. The measured/theoretical values of samples with high(21. 00 U/mL), medium(10. 50 U/mL) and low(1. 70 U/mL)concentrations were between 95% and 105%, with the RSDs within 15%. In the range of 0. 664-21. 263 U/mL, the R2was not less than 0. 99, the measured/theoretical values were between 95% and 105%, and the RSDs of precision verification was within 15%. The method had no difference in accuracy and precision at different incubation temperatures(35, 37, 39 ℃).The antigen content of CV-A6 antigen production process samples and finished products detected by the developed method effectively reflected the changing trend of antigen concentration in process samples and final products.Conclusion The double antibody sandwich ELISA for quantitative determination of CV-A6 antigen was developed with good linearity, range,specificity, accuracy, precision, and robustness, which can be used for the quality control of CV-A6 antigen process and final products, and lays a foundation for the in vitro efficacy evaluation of CV-A6 antigen.

In order to sum up the antineoplasmic active components of Holothuria, we studied the literatures in recent years. The results showed that Holothuria contained a variety of anti-tumor active ingredient, such as Holothurian Glycosaminoglycan and Holothurin. These anti-tumor active components played important roles in different stages of tumor developmment, progression and metastasis. The discovery of new anti-tumor active ingredients may be helpful for the application of Holothuria in clinical tumor treatment.

PBL teaching method can cultivate students’ ability to analyze the existing problems, find out more problems and resolve the problems finally, and it is important to improve students’ comprehensive quality. The experimental teaching innovation of Histology and Embryology is summarized in this article, which turned induction, practice and examination, the three stages of experimental teaching into" Problem-Based learning", thus making the teaching effect much better.

Ischemic cardiomyopathy (ICM) is due to coronary atherosclerosis cause long-term myocardial ischemia, make myocardial fibrosis diffuse, produce similar clinical syndrome like idiopathic dilated cardiomyopathy, which is one of a common clinical disease and causes increasing hazards to human health.Timely recovery of blood perfusion and coronary artery recanalization is one of the effective methods of treatment.However, it brings new damage while tissues and organs receive reperfusion, which is myocardial ischemia-reperfusion injury.In this paper, the current research progress in medicine myocardial ischemia-reperfusion injury and how to build a simple and efficient experimental animal models to mimic human myocardial ischemia-reperfusion injury were reviewed.

Objective To study the protective effect of the hypothermic brain-protection apparatus on hemorrhagic shock rabbits. Methods Applying the modified Wigger's shock animal model, we observed the effects of a self-developed brain-protecting apparatus on the survival time and vital signs of rabbits. Results Hypothermic brain protection could considerably reduce 60% lipid hyperoxide in brain tissues as compared with that in the control group. It could also decrease heart rate and respiration and hence reduce tissue oxygen consumption. Conclusion Hypothermic brain protection can attenuate the brain lesion and obviously prolong the survival time of hemorrhagic shock animals.

Objective To study the radiosensitization of histone deaeetylases inhibitor(HDACI) panobinistat in prostate cancer cells in vitro,as well as the possible mechanisms.Methods IC20 of two prostate cancer cell lines(LNCaP and PC-3)was determined using MTI assay.Cells received a single dose irradiation of 0,2,4,6,or 8 Gy using 6 MV X-ray for radiosensitivity experiment,but only 2 Gy for western blot and flow cytometry.Radiosensitization of panobinostat was investigated with clonogenic assay,and sensitizing enhancement ratio(SER)was calculated with single-hit multi-target model.Western blot was used to compare γH2AX expression.Flowcytomctry was used to detect the cell cycle distribution.Results IC20 of LNCaP and PC-3 was 2.5 and 10.0 μmol/L,respectively.SER of panobinostat at IC20 was 1.37(D0 ratio)and 1.11(Dq ratio)for LNCaP cells,and 1.78(D0 ratio)and 1.17(Dq ratio)for PC-3 cells.Expression of γH2AX gradually decreased in the 2 Gy irradiation-alone cells standing for the DSB repair,while γH2AX expression was persistent in the combination group.Irradiation triggered a G2/M arrest 6-12 hours after irradiation in LNCaP and PC-3 cells.G2/M arrest was observed when cells were treated with panobinostat for 24 hours,however,no significant change concerning cell cycle distribution was showed when cells received further irradiation.Conclusions Panobinostat Call radiosensitize prostate cancer cells,which may be related with increased DNA DSB,inhibition of DSB repair and attenuation of cell cycle modulation after irradiation.

Ischemic cerebrovascular disease(ICVD) is temporary or permanent reduction of blood flow of the main arteries of the brain caused by cerebral transient,sudden,reversible neurological dysfunction.Timely recovery of blood supply to the brain is the most important way to save the patient's life,but at the same time gave restore blood perfusion brain tissue organs brought new damage,which is cerebeal ischemia reperfusion injury(CIRI).How to build an ideal,efficient,easy to operate brain ischemia-reperfusion injury in animal models to simulate the clinical study,and how to improve and innovate this model by researchers in recent years were reviewed in this article.

OBJECTIVETo investigate the changes in serum amylase (AMY) and lipase (LPS) and their clinical significance in patients with acute paraquat poisoning (APP).

METHODSThe clinical data of 62 APP. patients who were admitted to Guangzhou 12th People's Hospital in 2014 were retrospectively analyzed. With clinical death being regarded as the end point of observation, the 62 patients were divided into survival group (n=32) and death group (n=30). The fasting venous blood of the patients on the 1st, 3rd, and 7th day after admission were collected, and the AMY and LPS levels were determined; the obtained data were compared between the two groups and analyzed statistically.

RESULTSThe AMY and LPS levels of the survival group on the 1st, 3rd, and 7th day after admission were significantly lower than those of the death group (P<0.05); the AMY and LPS levels of the death group on the 3rd day were significantly higher than those on the 1st day after admission (P<0.05); the survival group demonstrated no significant changes in AMY and LPS levels (P>0.05).

CONCLUSIONChanges in AMY and LPS levels in APP patients are reliable indicators for the determination of poisoning severity and prognosis.

Amylases ; blood ; Humans ; Lipase ; blood ; Paraquat ; poisoning ; Poisoning ; blood ; mortality ; Prognosis ; Retrospective Studies

Objective To explore the method of primary isolation, cultivation and identification of rat brain microvessel pericytes. Methods The brain tissue of 10 3 week-old Wistar rats was separated sterilely. The brain microvessel fragments were separated using two-step enzyme digestion and one-step gradient centrifugation and were seeded in 35-mm dishes for primary culture. The cell morphology was observed by phase contrast microscopy; the immunofluorescence assay was used to identify the associated antigns, such as the α-smooth muscle actin (α-SMA), neuron-glial antigen 2 (NG2), von Willebrand factor (vWF), and glial fibrillary acidic protein (GFAP). Methyl thiazolyl tetrazolium was used to determine the cell growth curve. Results Pericytes climbed out from the adherent brain microvascular fragments around,showing polygonal, and the cell fusion was 95％ after 12-14 days. Immunofluorescence staining revealed that the molecular markers of the pericytes α-SMA and NG2 related antigens showed double positive, while the vWF and GFAP related antigens showed double negative and the cultured cells were confirmed as brain microvascular pericytes. The growth rate of primary cells was slower. The passage cells entered into logarithmic growth phase after 36 to 60 hours and entered into plateau phase after 72 to 108 hours. Conclusions This method may successfully isolate rat brain microvascular pericytes with higher purity.

Objective To investigate the distribution and variation trend of whole-blood Ca,Cu,Zn,Fe and Mg concentrations of children in fengxian district,Shanghai,and provide a reference for prevention of elements deficiency in children.Methods 2 219 children in outpatient department of pediatrics were enrolled in the study,who reccieved the tests for five kinds of elements(Ca, Mg,Fe,Cu,Zn)in whole blood by using atomic absorption spectrometer.The children were divided into 5 age groups:infancy group (1-<12 months old),toddler age group(1 -<3 years old),preschool age group(3 -<7 years old),school age group(7 -<12 years old)and adolescence group(12-18 years old).The distributions and variation trend of the 5 kinds of elements were analyzed among different age and gender groups.Results The deficiencies of Ca,Fe and Zn were common in all the age groups,the total defi-ciency rates of Ca,Fe and Zn were 19.51%,22.08% and 25.82% respectively.there was no significant difference between boys and girls(P >0.05).Differences among different age groups were statistically significant(P <0.05).The Average concentrations of Zn displayed increasing tendency of different degrees in different groups of children,other elements displayed no obvious variation trend.Conclusion Children′s Zn,Fe and Ca deficiencies in Fengxian District,Shanghai are in different degrees,regular determina-tion of calcium and trace elements should be performed and take supplementary measures as soon as possible.