FTB多克隆抗体的研制

为建立ＦＴＢ的间接酶联免疫吸附方法，用自行制备的ＦＴＢＳ－载体蛋白结合物ＦＴＢＳ－ＢＳＡ和ＦＴＢＳ－ＨＳＡ作为免疫原，按常规免疫程序分别免疫新西兰大耳白家兔各２只，４～１０周后获得２种抗ＦＴＢ的多克隆抗体，ＰｃＡｂ－ＦＴＢ１（以ＦＴＢＳ－ＢＳＡ为免疫原）和ＰｃＡｂ－ＦＴＢ２（以ＦＴＢＳ－ＨＳＡ为免疫原）。以ＦＴＢＳ－ＩｇＧ作为标识抗原包被酶标板，建立了测定ＦＴＢ的间接酶联免疫吸附试验（ＩＣ－ＥＬＩＳＡ）方法，这２株多克隆抗体与８种真菌毒素的交叉反应均较小。

FTB单克隆抗体的研制与特性鉴定

为建立测定ＦＴＢ的ＩＣ－ＥＬＩＳＡ方法，用复合抗原ＦＴＢＳ－ＢＳＡ和ＦＴＢＳ－ＨＳＡ作为免疫原免疫ＢＡＬＢ／ｃ小鼠，取免疫小鼠脾细胞与鼠ＳＰ２／Ｏ－Ａｇ１４骨髓瘤细胞融合，经反复筛选，获得２株稳定分泌抗ＦＴＢ抗体的单克隆杂交瘤细胞株。免疫球蛋白亚型鉴定，１株为ＩｇＭ，１株为ＩｇＧ１。两株杂交瘤腹水的ＥＬＩＳＡ效价分别为１×１０－６和１×１０－８，用ＰＥＧ沉淀法和辛酸－饱和硫酸铵法对抗体腹水进行了纯化，并分别测定了２种单抗与ＦＴＢＳ－ＩｇＧ的亲和力常数以及与其它１１种真菌毒素的相对交叉反应，建立了测定ＦＴＢ的ＩＣ－ＥＬＩＳＡ法。

Construction and characterization of an experimental ISCOMS-based hepatitis B polypeptide vaccine

AIM: To characterize the biochemical and immunological properties of an experimental ISCOMS vaccine prepared from a novel therapeutic polypeptide based on T cell epitopes of HBsAg, and a heptatis B-ISCOMS was prepared and investigated. METHODS: An immunostimulating complexes(ISCOMS)-based vaccine containing a novel therapeutic hepatitis B polypeptide was prepared by dialysis method, and its formation was visualized by electron microscopy and biochemically verified by SDS-polyacrylamide gel electrophoresis. Amount of the peptide within ISCOMS was determined by Bradford assay, and specific CTL response was detected by ELISPOT assay. RESULTS: Typical cage-like structures of submicroparticle with a diameter of about 40nm were observed by electron microscopy. Results from Bradford assay showed that the level of peptide incorporation was about 0.33g.L(-1). At the paralleled position close to the sixth band of the molecular weight marker(3480kDa) a clear band was shown in SDS-PAGE analysis, indicating successful incorporation of polypeptide into ISCOMS. It is suggested that ISCOMS delivery system could efficiently improve the immunogenicity of polypeptide and elicit specific immune responses in vivo by the results of ELISPOT assay, which showed that IFN-gamma producing cells(specific CTL responses) were increased(spots of ISCOMS-treated group: 47+/-5, n =3; control group: 5+/-2, n =3). CONCLUSION: ISCOMS-based hepatitis B polypeptide vaccine is successfully constructed and it induces a higher CTL response compared with short polypeptides vaccine in vivo.

Humoral and cellular immunogenecity of DNA vaccine based on hepatitis B core gene in rhesus monkeys

INTRODUCTIONHepatitis B virus (HBV) is the most commonetiologic agent for infectious liver diseases. It isestimated that there are more than 250 millionchronic HBV carriersin the world today and thereis a significant association among persistentinfection, liver cirrhosis and hepatocellularcarcinoma[1-3].

A comparative study on serologic profiles of virus hepatitis B

INTRODUCTIONHepatitis B viral infection, one of the most-prevalent liver disorders in China and Korea, is aserious infectious disease as it has the potential ofprogressing into liver cirrhosis and primary hepaticcarcinoma. China and Korea both belong to high-risk endemic regions of viral hepatitis[1]. TheHBsAg positive rates in China ranged from 6.9% -17.9% by age, race and test methods[2-5].

Basic Study Hepatitis B virus detected in paper currencies in a densely populated city of India: A plausible source of horizontal transmission?

BACKGROUND The recent rise in the incidence of hepatitis B virus(HBV)infections in a densely populated city of eastern India(“mixing vessel”of people of varied socioeconomic and immune status)prompted this study.Applying saliva on fingers for enumerating bank notes is a common practice in the Indian subcontinent.Paper notes may be a potential source of“horizontal”transmission of this virus,especially if there are cuts/bruises on the oral mucous membrane or skin.AIM To investigate whether paper currencies could be a plausible mode of horizontal transmission of HBV infection.METHODS Polymerase chain reactions(PCR)followed by nucleotide sequencing was done for the detection of HBV.Hepatitis B virus surface antigen enzyme-linked immunosorbent assay(HBsAg ELISA)was performed on all HBV deoxyribonucleic acid-positive samples to check the detectability of the virus.Atomic force microscopy(AFM)was carried out for visual confirmation of HBV particles in ultracentrifuged/immunoprecipitated samples from currency paper washings.RESULTS HBV-specific PCRs on pellets obtained after ultracentrifugation/immunoprecipitation of the currency paper washings detected potentially intact/viable HBV(genotype D2)in 7.14%of samples(n=70).AFM gave the visual confirmation of HBV particles in ultracentrifuged/immunoprecipitated samples from currency paper washings.However,HBV isolates from the currency notes could not be detected by HBsAg ELISA.CONCLUSION It is a common practice in the Indian subcontinent to count paper currencies by applying saliva on fingertips.Paper notes may be a potential source of“horizontal”transmission of this virus,especially if there are cuts/bruises on the oral mucous membrane or skin,but it was practically not possible to demonstrate experimentally such transmission.Detection of potentially intact/viable and“occult”HBV from currency poses potential risk of silent transmission of this virus among the general population.

Dynamic change of serum protein S100b and its clinical significance in patients with traumatic brain injury

Objective: To analyze the dynamic change of serum protein S100b in patients with traumatic brain injury and its clinical value in assessing brain damage. Methods: According to Glasgow coma scale (GCS), 102 cases of traumatic brain injury were divided into mild brain injury group (GCS≥13, n=31, Group A), moderate brain injury group (8<GCS<13, n=37, Group B) and severe brain injury group (GCS≤8, n=34, Group C). Serial S100b concentrations were analyzed by enzyme-linked immunosorbent assay (ELISA) in blood samples taken on admission, 12 h, 24 h, 48 h, 72 h and 7 days after traumatic brain injury. Results: The severe brain injury group showed significantly higher concentration of serum S100b, with earlier increase and longer duration, than the mild and moderate brain injury groups. The patients with higher S100b exhibited lower GCS scores and poor clinical prognosis. The increase in S100b could emerge before clinical image evidence indicated so. Conclusions: Serum S100b can be used as a sensitive index for assessment and prediction of traumatic brain injury severity and prognosis.

Analysis of aflatoxin B1 in contaminated feed,media,and serum samples of Cyprinus carpio L.by high-performance liquid chromatography

Objectives:Enzyme-linked immunosorbent assay(ELISA)-,thin-layer chromatography(TLC)-,and highperformance liquid chromatography(HPLC)-sensitive methods were used for the identification of aflatoxin B1(AFB1)contaminated fish feed,media,and fish serum samples.Materials and Methods:The ELISA,TLC,and HPLC methods were validated by the quantitative and qualitative determination of AFB1 in contaminated fish feed,media,and fish blood serum samples.Results:The primary identification of AFB1 was carried out with a DOA-ELISA test kit ELISA followed by TLC with RF values 0.81,0.79,0.81,and 0.80 of AFB1-contaminated fish feed,media,and serum samples,respectively,compared with AFB1 standard.HPLC results show that the AFB1 levels in contaminated fish feed,media,and serum samples were 2.6,2.6,and 2.7 ng/mL,concentrations respectively.The level of concentrations of AFB1 was almost similar in all three samples,but slightly higher in the fish serum sample with,2.7 ng/ml.However,the present method is strongly recommended for monitoring AFB1 contamination in feed stuffs,especially in fisheries where the feed is under continuous exposure to moisture.Conclusions:This method is accurate and more sensitive when compared with routine conventional AFB1 detection methods,and is highly applicable in aquaculture and fisheries to screen the mycotoxins in fish feed.