分子生物学在鱼类分类上的应用

<正> 近二十五年来,分子生物学取得了惊人的发展,成为今天生物学研究的径流,它已渗入和影响了生物学的各个领域,产生了一批新兴学科,如分子遗传学、分子细胞学、分子分类学、分子神经生理学、分子药理学、分子病理学等.甚至传统的鱼类分类学也     

仔稚鱼磷脂营养需求的研究进展

磷脂(Phospholipid,PL)又被称为"极性脂",是指分子中含有磷酸的复合脂,在自然界分布广泛,所有的细胞中都含有磷脂.目前磷脂在世界上已备受人们的青睐,有关磷脂的研究工作也已深入到各个领域,在世界范围内的食品、保健品、医药以及饲料行业得到越来越广泛使用.     

表面活性剂提高养殖水体溶氧量原理初探

表面活性剂品种繁多,商品牌号已达20000多个,产量达1200万吨,其用途非常广泛,涉及工农业及人民生活的各个领域,有"工业味精"的美称.而其中应用于水产养殖的很少,至于有关表面活性剂在水产养殖中的应用论文就更少了.     

锯缘青蟹凝集素的提取及单克隆抗体制备

凝集素是无脊椎动物非特异性免疫系统的重要组成成份,可凝集外来病原菌、进行非己识别、调理和介导血细胞吞噬。开展了锯缘青蟹血清凝集素的提取、单克隆抗体制备及凝集素定位研究,以N-乙酰基葡萄糖胺（N-Acetyl-D-glucosamine,GlcNac）为配基制备了Sepharose 4B亲和层析柱,可选择性结合青蟹血清凝集素;青蟹血清通过亲和层析柱后,以0.45 mol/L NaCl洗脱,得到有较高凝集活性的单一洗脱峰,该洗脱峰凝集素比活为血清凝集活性的32.6倍,SDSPAGE显示存在87 ku和79 ku主要蛋白成分;以纯化青蟹凝集素免疫BALB/c小鼠,经细胞融合、单抗筛选获得BF82等7株可稳定分泌抗凝集素单抗的小鼠杂交瘤细胞株,单抗腹水的ELISA效价为1∶10⁴～1∶10⁵;单抗可特异性抑制青蟹凝集素的血凝,血凝抑制效价为1∶32～1∶64,亚型鉴定表明7株单抗均为IgG1型,免疫印迹表明单抗可特异性结合87 ku、79 ku蛋白;采用凝集素单抗间接免疫荧光法进行了青蟹血淋巴细胞染色,结果显示凝集素主要分布于颗粒细胞的细胞质颗粒和透明细胞的细胞膜上。     

国外鱼类疫苗之路

疫苗接种在世界范围内鱼类大规模商业养殖中的作用日益显著,并已成为国外以鲑鱼为代表的重要经济鱼类养殖业成功的关键因素之一.目前,国际水产养殖市场上主要养殖品种,如鲑鱼、鳟鱼、欧洲鲈鱼、鲶鱼、大西洋鳕鱼等,都有了相应的商品化疫苗,接种疫苗已成为当前国际水产养殖业的规范性生产标准.     

欧洲鳗黏膜免疫球蛋白单克隆抗体的制备及其特性分析

应用杂交瘤单克隆抗体技术制备了8个分泌抗欧洲鳗（Anguilla anguilla）黏膜免疫球蛋白的单克隆抗体细胞株，并对这些单克隆抗体的特性进行了分析。结果显示，8株单抗中IgM有2株，IgG，和IgG2。各有3株；所有单抗均与欧洲鳗黏膜免疫球蛋白呈ELISA反应阳性，腹水抗体滴度为在10^4～10^6之间。进一步实验证实，这些单抗与供试的10种水产动物常见病原菌均无交叉反应；其中3株单抗与欧洲鳗血清IgM有不同程度的交叉反应；4株单抗与黏膜免疫球蛋白有交叉反应，其中2株单抗交叉反应较弱。以上结果提示，欧洲鳗黏膜免疫球蛋白和血清IgM之间既有各自独特的抗原决定簇，又有共同抗原位点，证实欧洲鳗黏膜抗体在抗原性方面有别于血清IgM。成功制备的这8株单抗可用于鳗黏膜免疫球蛋白的检测及其结构和功能分析，为欧洲鳗黏膜免疫的进一步研究提供工具。[中国水产科学，2008，15（3）：511-515]     

转基因技术在动物营养学中的应用及发展前景

转基因技术的研究和应用已渗透到生命科学的各个领域,动物营养学的发展需要在分子水平上分析及解释营养物质对动物机体的变化调控,如生长发育、新陈代谢、遗传变异、免疫与疾病等。本文综述了转基因动物在动物营养学中的应用:改善生产性状,提高生产性能;建立遗传性疾病、肿瘤和其它疾病的实验动物模型;增强抗病力;利用转基因动物生产药用蛋白质。     

广东省两行政部门资源整合全力发展滨海旅游 滨海旅游流量来了!

正在资本场上,每过一段时间,线上流量价格就上跳一个数字,视流量为生命线的创业者们和坐拥流量的少数巨头在此间博弈,而无数普通用户被反复细分在各个领域后再次汇聚在一起,成为了资本市场上不容小觑的力量。只是,这与政府发展滨海旅游有什么关系?3月7日,广东省海洋与渔业厅与广东省旅游局签订的合作框架协议,就是吸引流量的举措之一。这种资源整合在不少业内人士看来,非常有必要,也是推动广东滨     

卫星技术在海洋及渔业上的应用

半个世纪以来,航天科技与卫星技术突飞猛进,已广泛应用到交通、资源、环境、海洋、科研、军事等各个领域。海洋渔业是最重要的传统海洋产业之一,当前,海洋渔业的可持续发展面临着资源衰退、环境污染、渔船监控与管理等亟需解决的问题,卫星技术在海洋及渔业上的应用可为解决这些问题提供有效的技术支撑,并已成功得到若干应用。     

龟鳖产品品牌的创立与保护

我国是世界上最大的龟鳖养殖和加工国家,龟鳖已成为农村致富的特种养殖品种之一.虽然我国龟鳖产业在发展的过程中经过了价格起落、盲目发展等风雨,但优质龟鳖产品已越来越受国内外消费者的欢迎.所以生产安全放心的优质产品,走龟鳖产品精品之路是今后发展龟鳖产业的方向.为此,笔者就龟鳖产品怎样树品牌、创名牌,谈几点自己的经验,供同行参考.     

Biological characterization of a monoclonal antibody against a surface membrane antigen on Cryptobia salmositica Katz, 1951

A monoclonal antibody (IgG 1) (designated as MAb-001) was produced against the pathogenic haemoflagellate Cryptobia salmontica Katz. The antibody agglutinated live parasites under in vitro conditions. Live C. salmositica, incubated with MAb-001 at 10 °C, did not multiply and were dead within 4 weeks in culture. About 50% of juvenile rainbow trout, Oncorhynchus mykiss (Walbaum), inoculated intraperhoneally with C. salmositica, incubated in MAb-001 prior to inoculation, did not become infected, while in adult rainbow trout, the peak parasitaemia was reduced. These results indicate that MAb-001 is a protective monoclonal antibody and the antigen it recognizes is located on rhe surface membrane of C. salmositica. The antibody also inhibits multiplication and affects viability of the parasite under in vitro conditions.     

Monoclonal antibody-based immunodot for evaluating serum immunoglobulin levels in rohu (Labeo rohita)

Immune response in rohu treated with an immunomodulator is usually evaluated employing either non-specific immune parameters or traditional antibody-based tools. In the present study, a monoclonal antibody-based immunodot has been developed for evaluating antibody titre in rohu as a preliminary tool to ensure antibody response due to the effect of an immunomodulator, which can be used for routine field level analysis. The immunodot was sensitive enough to determine rohu immunoglobulin up to 15 μg/ml. Application of the immunodot for evaluating enhancement in immune response could be successfully demonstrated in probiotic fed rohu.     

抗犬瘟热高免卵黄抗体IHA检测方法的建立

目前检测卵黄抗体的方法很多,但IHA方法具有简便快速的优点,本研究的目的是为了建立抗犬瘟热高免卵黄抗体检测方法。采用犬瘟热病毒和犬细小病毒二联疫苗处理作为疫苗抗原,用犬瘟热病死犬的肝脏部分处理后作为组织抗原,制备敏化绵羊红细胞,用IHA方法确定了犬二联疫苗抗原和犬瘟热病毒组织抗原的最适含量。二联疫苗提取抗原的含量为0.7 mg/mL致敏红细胞时,与犬瘟热单克隆抗体反应显著,抗体效价为1∶256;而患犬瘟热松狮犬的肝脏组织抗原蛋白浓度为0.5mg/mL,与犬瘟热单克隆抗体反应显著,抗体效价为1∶128。经过IHA验证,制备的松狮犬瘟热肝组织抗原,蛋白含量0.5 mg/mL 1%致敏红细胞,对应的单抗、鸡血清均出现了凝集,阳性对照也出现凝集,阴性对照未出现凝集。建立的IHA抗体检测方法,为准确检测制备抗犬瘟热高免卵黄抗体的效价奠定了基础。     

Use of monoclonal antibodies in the indirect fluorescent antibody technique (IFA) for the diagnosis of Edwardsiella ictaluri*

Abstract. Forty-five channel catfish, Ictalurus pitnctaius (Rafinesque), fingerlings were inoculated with Edwardsiella ictaluri to determine the usefulness of monoclonal antibodies for indirect fluorescent antibody techniques for confirming clinical diagnosis of enteric septicaemia of catfish. Bacteriological and indirect fluorescent antibody results were compared statistically and found to correlate in 90·3% of the cases. A method for incorporating monoclonals for indirect fluorescent antibody into the speciation of Edwardsiella ictaluri and the use of monoclonal antibodies in diagnosis is described.     

血卵涡鞭虫单克隆抗体的制备与初步应用

用血卵涡鞭虫可溶性抗原免疫BALB/c小鼠,经常规融合、间接ELISA方法筛选,将所得阳性克隆再经3次亚克隆后,共获得3株针对血卵涡鞭虫的单克隆抗体(2B₂、3G₄、4G₇),单克隆抗体亚类鉴定表明,三者为IgG类抗体。用筛选的杂交瘤细胞株制备小鼠腹水抗体,其细胞上清及腹水效价分别为5.12×10^-4和8.00×10^-4。进一步利用单克隆抗体建立间接荧光抗体方法对单抗特异性进行鉴定,阳性虫体被染上黄绿色荧光,而正常梭子蟹血淋巴则未被染色。用单克隆抗体和多克隆兔抗血清以羊抗鼠HRP-IgG为酶标抗体,建立了检测血卵涡鞭虫的双抗体夹心ELISA方法,该方法对血卵涡鞭虫阳性标本检测符合率为100%。结果表明,制备的单克隆抗体效价高、特异性好,可用于血卵涡鞭虫的早期临床诊断。     

Rapid development of polyclonal antisera against infectious salmon anaemia virus and its optimization and application as a diagnostic tool

Infectious salmon anaemia is an important disease of Atlantic salmon. One of the current methods of diagnosis is the indirect fluorescent antibody test (IFAT), using a monoclonal antibody specific to the haemagglutinin of the virus. The conformationally dependent nature of this antibody could be a drawback in its usefulness in other tests. This study describes the development and optimization of a polyclonal antiserum against infectious salmon anaemia virus, including a method of separating virus from cell culture components within culture supernatant. The antiserum was subsequently optimized for use in a variety of immunological diagnostic tests, including IFAT and an alkaline phosphatase-based immunoassay, and Western blot.     

Characterization of a monoclonal antibody against the 47- kDa antigen of Cryptobia salmositica Katz and its use in an antigen-capture enzyme-linked immunosorbent assay for detection of parasite antigen in infected rainbow trout, Oncorhynchus mykiss (Walbaum)

A monoclonal antibody (designated MAb-007) was produced against the pathogenic haemoflagellate Cryptobia salmositica Katz. This IgG3 antibody recognized the 47-kDa antigenic polypeptide of C. salmositica (SDS-PAGE and Western immuno-blotting). The antibody did not agglutinate live parasites, and there was no change in the staining intensity of the 47-kDa band on Western immunoblots after immunoabsorption of MAb-007 with live intact parasites. The 47-kDa antigen recognized by MAb-007 was localized in the cytoplasm of the parasite (immunogold labelling and electron microscopy). The monoclonal antibody cross- reacted with the 47-kDa polypeptides of C. bullocki Srrout and C. catostomi Bower & Woo. It was used in an antigen-capture ELISA for the detection of parasite antigen in the plasma of rainbow trout inoculated with the parasite, or with an attenuated vaccine strain of C. salmositica. All pre-infection plasma were negative while all infected fish with detectable parasitaemias were positive for antigen at 1–9 weeks after infection. Parasite antigen was even detected in vaccinated fish that were negative for parasites using the wet mount microscopic technique. The antigen-capture ELISA detected C, salmositica antigen in whole cell lysate preparations at concentrations as low as 0.5 μg ml^-1. Fifty microlitres of fish plasma was required in the antigen-capture ELISA, and the use of a plate reader and 96-well plates facilitated rapid analysis of a large number of plasma samples. The sensitivity of the assay makes it a potentially useful tool for detection of Cryptobia infections.     

Detection methods of Cyprinid herpesvirus 2 infection in silver crucian carp (Carassius auratus gibelio) via a pORF72 monoclonal antibody

Cyprinid herpesvirus 2 (CyHV‐2) is the main pathogen responsible for causing haematopoietic necrosis disease in Carassius auratus gibelio. Although many nucleic acid‐based diagnostic methods have been applied, no stable and sensitive immunological diagnostic approaches have been reported. In this study, to detect CyHV‐2 in clinical samples using immunological methods, recombinant ORF72 protein (pORF72), encoded by the CyHV‐2 ORF72 gene, was used as a capture antigen to identify blood and tissues infected with CyHV‐2. First, ORF72 gene was amplified from the CyHV‐2 genome and cloned into a PGEX‐4t‐3 expression vector to produce pORF72 in Escherichia coli. The purified pORF72 was used as an immunogen to prepare monoclonal antibodies. The Western blotting assays revealed that the monoclonal antibody could specifically identify the pORF72. Furthermore, an immunohistochemical protocol and a blood smear method were established to detect CyHV‐2 in carps. The results indicate that the monoclonal antibody against pORF72 could be utilized as an effective detection tool for haematopoietic necrosis disease in Carassius auratus gibelio.     

A monoclonal antibody cross-reactive with three salmonid herpesviruses

Abstract. A monoclonal antibody cross-reactive with strain H-83 Oncorhynchus masou virus (OMV) and Herpesvirus salmonis has been prepared. Western blot analysis revealed that the monoclonal antibody, designated clo. 7, recognized the 16 000 (16K)-polypeptide of these herpesviruses but not the polypeptides of infectious pancreatic necrosis virus or infectious haematopoietic necrosis virus. Clo. 7 showed neither neutralizing activity against strain H-83 nor cytotoxicity to strain H-83-infected RTG-2 cells in the presence of complement. The purified antigen, recognized by clo. 7, from strain H-83 was a polypeptide with a molecular weight of 16K, and sensitive to proteolytic enzymes and sodium periodate, but not to neuraminidase and ethylether. The antigen was heat stable and stable at pH 5·0 – 9·0.     

Immunoglobulin heterogeneity in the rainbow trout, Salmo gairdneri Richardson

Abstract. Hetereogeneity of rainbow trout immunoglobulins was demonstrated by using monoclonal antibody 1A6 and polyacrylamide-gradient gel electrophoresis. Immunoglobulins defined by elisa using monoclonal antibody 1A6 were about 30% of the total immunoglobulins, detected by elisa using polyclonal antibodies, in healthy rainbow trout. In trout obtained from farms with a previous history of infectious viral diseases, 1A6-immunoglobulins were only about 14% of the total. Several serum pools from infected trout could be totally depleted of 1A6-immunoglobulins (about 12% of total immunoglobulins) by affinity chromatography over Sepharose immobilized monoclonal antibody 1A6. Polyacrylamide-gradient gel electrophoresis under denaturing conditions of total immunoglobulins, 1A6 immunoglobulins and no-1A6 immunoglobulins purified by affinity chromatography, showed a majority heavy chain of 70 KDa and a minority heavy chain of about 60 KDa, two light chains of 24 and 26 KDa, and a 11–14 KDa polypeptide.