脱落酸抗体的抗独特型抗体的制备与鉴定

用ABA多克隆抗体(Ab_1)以金黄色葡萄球菌菌体(SPA)作为载体,免疫家兔,制备的抗独特型抗体(Ab_2)初步纯化后用ELISA试验鉴定其特异性的结果表明,该抗独特型抗体具有良好的特异性,能阻断ABA抗体对ABA的结合反应,并与ABA结合蛋白结合,暗示其有模拟抗原的作用。 Jerne的免疫网络学说认为,特异性抗体(Ab_1)可变区中的独特型决定簇(Id)可诱导抗独特型抗体(Ab_2)的产生,Ab_2中的一部分可以模拟抗原的作用。Sege和Peterson提出,     

抗肺炎支原体卵黄抗体的制备及其免疫特性

目的制备抗肺炎支原体卵黄抗体,并研究其免疫特异性。方法以超声粉碎法制备肺炎支原体抗原;以ELISA法测定卵黄抗体的效价及免疫特异性;以水稀释法联合疏水层析的方法分离纯化卵黄抗体;应用SDS-PAGE法测定分子量及鉴定抗体纯度;改良Lowry法测定蛋白含量。结果低、高剂量组均诱导母鸡产生有效免疫应答,高剂量组免疫效价高于低剂量组。高剂量组于初免疫后约50d抗体效价达高峰,持续约2个月;而低剂量组在初免疫后约60d抗体效价达高峰,持续约1个月。之后效价逐渐下降,在免疫约120d,高剂量组由13log2下降到10log2;而低剂量组则由11log2下降到7log2。以水稀释法联合疏水层析法制备了电泳纯抗肺炎支原体IgY,分子量约178KD,平均每1ml卵黄液可获得较纯抗体6.4mg。制备的IgY与肺炎支原体具有较高特异性,与解脲支原体和人型支原体无明显交叉反应,与生殖支原体有轻度的交叉反应。结论本研究初步制备了抗肺炎支原体卵黄抗体,为肺炎支原体的防治与检测提供新的途径。     

抗α-INF卵黄抗体的制备与鉴定

用人抑制素（１－２６）Ｔｙｒ．Ｇｌｙ与ＫＬＨ连接作免疫原,免疫本地良种母鸡,观察血清及卵黄中特异性抗体的应答,结果经免疫的鸡免疫应答,所产卵的卵黄中可检出大量的有高度活性的抗体。综合运用ｐＨ值、非示蛋白沉淀法和球蛋白沉淀法,制得纯度较高的抗抑制素ｅＩｇＹ。采用阻断ＥＬＩＳＡ法证实所获得的ｅＩｇＹ具有高度的特异性。用ｅＩｇＹ被动免疫未成年昆明鼠,子宫重量明显增加,由此可以用鸡做生物反应器大量生产抗抑     

抗独特型抗体研究的新进展

独特型是存在于抗体分子某些部位上的一种特殊基因。该基因具有抗原性,能在自体或异体内诱导出针对该基因的特异性抗体——抗独特型抗体。近年大量的动物实验证明,抗独特型抗体可以作为新一代免疫制剂,用以弥补现有疫苗的不足。同时,它还可用于癌症和自身免疫性疾病的治疗。     

布鲁氏菌抗独特型抗体的研究

在建立布鲁氏菌单克隆抗体细胞株的基础上,筛选具有保护作用的单抗A7免疫家免制备抗独特型抗体(简称二抗)。经阻断试验和竞争抑制试验证实,该二抗具有抗原“内影象”。将其提纯后配制成甘油佐剂苗免疫豚鼠和小白鼠．检测结果表明：免疫豚鼠和小白鼠产生了布鲁氏菌凝集抗体;免疫小鼠的循环T ANAE-细胞百分数明显高于对照组;免疫豚鼠在免疫后4个月时强毒菌株攻击有79．2％获得保护。因此认为,该二抗不仅具有抗原的。内影象”结构,而且具有良好的免疫原性,可供作疫苗用。     

噬菌体呈现技术制备结肠癌抗独特型抗体

 以纯化的鼠抗人结肠癌细胞单克隆抗体 (简称单抗 )MC5与钥孔血蓝素 (KLH)的交联物经腹腔免疫Balb c小鼠 ,取脾分离mRNA .RT PCR分别扩增抗体重、轻链可变区基因片段 (VH 和VLcD NA ,大小分别约为 340bp和 32 0bp) ,二者经linkerDNA连接形成ScFv(singlechainvariablefragment)DNA(约 75 0bp) .将ScFvDNA与噬菌粒载体pCANTAB5E的连接产物转化于大肠杆菌TG1,经辅助噬菌体M13KO7感染后 ,获得重组噬菌体抗体ScFv文库 .以单抗MC5对ScFv文库进行 4轮亲和筛选后 ,随机挑取 80个克隆经酶联免疫吸附实验 (ELISA)筛选出 2 2个呈现ScFv形式抗独特型抗体(抗 IdScFv)的噬菌体单克隆 .竞争抑制实验表明 ,在 2 2个阳性克隆中有 4个克隆所呈现的抗 IdScFv属β或γ型 .针对单抗MC5的噬菌体呈现型抗 IdScFv的制备 ,为筛选新的结肠癌重组抗 Id瘤苗候选分子奠定了基础     

糖类抗原的抗独特型抗体疫苗

糖类抗原以其免疫性较弱,常规疫苗的免疫效果一般欠佳,特别是对新生儿的免疫更是如此。糖类抗原又以其本身结构上的复杂人工合成较为困难,也不能通过重组DNA技术由基因工程直接制得。为此,不少免疫学家都在探寻有效的抗糖疫苗。本文作者长期从事糖类抗原的抗位特型抗体研究,认为目前的研究为寻求一条糖类抗原的抗独特型抗体疫苗奠定了基础;列举了细菌的果聚糖,大肠杆菌K13多糖,克氏锥虫的表面糖蛋白, A组链球菌糖,脑膜炎奈瑟氏菌C组多糖,以及肿瘤相关抗原上的糖表位等的研究材料。本文为1991年出版的抗独特型抗体疫苗一书中的一章。     

Preparation and application of anti-idiotypic antibody against anti-gibberellin A4 antibody.

A monoclonal anti-idiotypic antibody was raised against anti-gibberellin A4 (GA4) antibody, which recognizes biologically active gibberellins such as GA1 and GA4 specifically. Amino acid sequences of variable regions of both anti-GA4 and anti-idiotypic antibodies were analyzed. By using the property of the anti-idiotypic antibody to compete with GA1/4 in binding to the anti-GA4 antibody, we successfully applied the anti-idiotypic antibody to ELISA as a tracer for measuring GA1/4. The single-chain Fv (scFv) gene of the anti-idiotypic antibody was constructed, and scFv expressed in E. coli showed binding activity to anti-GA4 antibody. These results suggest the possible application of anti-idiotypic antibody as a handy and stable source of an enzymatic tracer for ELISA by production of fusion protein of the scFv and an appropriate enzyme.     

Biologic activity of anti-thyrotropin anti-idiotypic antibody

We raised an antihuman thyrotropin anti-idiotypic antibody and showed that it was active at the thyrotropin receptor. Thus this antibody inhibited ¹²⁵I b-TSH binding to thyroid plasma membranes, stimulated adenylate cyclase activity through a guanyl nucleotide-dependent mechanism, increased radioiodide entry rate into isolated porcine thyroid follicular cells, and induced such cultured cells to organize into follicles. All these parameters are typical of thyrotropin action. This work raises the possibility that thyroid stimulating antibodies that cause the hyperthyroidism of Graves disease may be, at least in some patients, anti-thyrotropin anti-idiotypic antibodies. It also offers a novel method whereby antireceptor antibodies used in the isolation and characterization of the receptor may be raised from ligands.     

Preparation of an anti-idiotypic antibody directed against anti-dexamethasone antibody and demonstration of its ability to acknowledge the receptor of glucocorticoids

By using a 3-carboxymethyloxime dexamethasone derivative coupled to bovine serum albumin we have prepared specific anti-dexamethasone antibodies in rabbits. These antibodies were then purified by affinity chromatography and administered to a second set of rabbits. One of them produced anti-idiotypic antibodies able to impede the [3H] dexamethasone binding of the initial anti-dexamethasone antibodies and to displace the [3H] dexamethasone-antibodies complexes towards high molecular weight species in gel filtration experiments. Moreover these antibodies we were also able to impede the binding of [3H] dexamethasone to the rat liver glucocorticoid receptor and to recognize the highly purified receptor using Western blot analysis.     

抗独特型抗体与T细胞记忆的关系

记忆T细胞作为人体免疫系统中的一个组成部分,在免疫应答中发挥着至关重要的作用,因此利用抗独特型抗体制备诱导产生记忆T细胞的疫苗是免疫学领域的一个重要方向。抗独特型抗体Fab段具有与特异性抗原相似的抗原决定簇的结构,其作为抗原替代物制备的疫苗所激发机体产生的记忆T细胞具有特异性强和安全性高的特点,成为一种比较理想的疫苗.就抗独特型抗体与T细胞记忆之间的联系及其应用效果作一简要综述。     

An antibody and anti-idiotypic antibody against the extra signal peptide of pre-aspartate aminotransferase

A peptide (extra signal peptide) comprising amino acids 1-29 of pig liver pre-mitochondrial aspartate aminotransferase (p-mAAT) was synthesized chemically. The peptide was found to block the import of rat liver p-mAAT into rat liver mitochondria. An antibody raised against the peptide immunoprecipitated rat liver p-mAAT synthesized in a rabbit reticulocyte cell-free translation system. These results suggested that the extra signal peptide sequence of p-mAAT is essential for import of p-mAAT into the mitochondria and that there is structural homology between the extra signal peptides of pig and rat liver p-mAAT. An anti-idiotypic antibody against the peptide was also prepared and purified by affinity chromatography on an Affi-Gel 10 anti-peptide IgG column and was then characterized.     

抗人血栓调节蛋白单克隆抗体的制备与鉴定

为了制备特异性抗人血栓调节蛋白(human thrombomodulin,hTM)的单克隆抗体(monoclonal antibody,McAb),利用脂质体Lipofectamine 2000将包含hTM全长cDNA序列的重组表达质粒pThr402转染CHO细胞,经G418筛选及相关鉴定后获得高效稳定表达hTM的CHO-TM5细胞株。将CHO-TM5细胞直接免疫Balb/c小鼠,应用杂交瘤技术,通过细胞ELISA (cellular enzyme-linked immunoabsorbent assay,CELISA)筛选出阳性克隆后,将杂交瘤细胞株腹腔注射Balb/c小鼠诱生腹水。用CELISA、流式细胞术、免疫组织化学染色法及免疫印迹法对所获McAb的特异性进行鉴定。我们获得了1株可稳定分泌抗hTM的McAb的杂交瘤细胞株NH-1,其亚型为IgGl,McAb腹水效价为1×10~(-6),腹水抗体含量为20 mg/ml。NH-1对相应抗原具有较高的组织特异性,在体内与正常组织的交叉反应少,对人脐静脉内皮细胞、CHO-TM5有特异性结合反应,说明NH-1可特异性识别天然的hTM分子,为进一步应用此McAb进行hTM生物学功能及临床意义研究提供了基础。     

Interferon-like activity in an anti-interferon anti-idiotypic hybridoma antibody

We describe here an anti-idiotypic hybridoma antibody directed against affinity-purified rabbit idiotypic antibodies (Rb-Id) to a homogeneous protein, the recombinant human leukocyte A interferon (rIFN-alpha A). The supernatant of the hybridoma, designated 3-1B, was able to inhibit the neutralization of rIFN-alpha A activity by the idiotypic antibodies. An in vivo passage of uncloned 3-1B cells yielded hybridoma cells (presumably a subclone), designated 3B1, the supernatant of which exhibited interferon-like antiviral activity with both bovine kidney (MDBK) cells and human amnion (WISH) cells. This activity could be absorbed by polymer-bound goat anti-mouse immunoglobulin serum and by Rb-Id coupled to Affi-Gel 10, and could be partially eluted from the latter at pH 2.5. The anti-idiotypic hybridoma antibody was able to compete with 125I-rIFN-alpha A for binding to the Rb-Id and also to interferon receptor-bearing MDBK cells. The clinical significance of an interferon-like anti-idiotypic antibody is discussed.     

Fulminant Mycoplasma pneumoniae pneumonia.

The frequency of fulminant pneumonia due to Mycoplasma pneumoniae is relatively rare despite the high prevalence of Mycoplasma species infection in the general population. We recently encountered such a case and have reviewed the English-language literature on cases of M pneumoniae pneumonia that have resulted in respiratory failure or death. Due to host factors or on epidemiologic grounds, fulminant cases seem to be more common in young healthy adults, in males, and possibly in smokers among the 46 patients we found. An enhanced host cellular immune response may be responsible for the development of severe cases. A spectrum of small airways disease is characteristic, including cellular bronchiolitis and bronchiolitis obliterans with and without organizing pneumonia. Based largely on anecdotal experience, corticosteroid use may be salutary in patients with respiratory failure. For reasons that are not well known, the incidence of pulmonary thromboembolism is increased in fatal cases.     

Detection and identification of anti-idiotypic antibodies in heterogeneous immunoenzyme analysis

The antigen-antibody equilibrium in the presence of a competitive agent of immunoglobulin nature, represented by anti-idiotypic antibodies, has been analyzed in the case of the immobilized antigen. As shown in this investigation, the amount of immunoglobulin bound with the carrier is determined both by the competition of anti-idiotypic antibodies for the active center of idiotype-carrying antibodies and by the binding of anti-idiotypic antibodies with the antigen-antibody complex. If idiotype-carrying and anti-idiotypic antibodies are represented by serum immunoglobulin of the same species, the titration of the competitive agent leads to obtaining a characteristic relationship between the amount of immunoglobulin bound with the insoluble phase and the amount of the competitive agent in the system passing through the maximum. The theoretical results have been confirmed on the experimental model and used for the identification of auto-antiidiotypic antibodies in patients' sera.     

Prevention of murine cryoglobulinemia and associated pathology by monoclonal anti-idiotypic antibody

A murine IgG3 mAb, clone 6-19, derived from non-manipulated autoimmune MRL/MpJ-lpr/lpr mice is a rheumatoid factor specific for IgG2a and is able to generate cryoglobulins via nonspecific IgG3 Fc-Fc interaction. Intraperitoneal passive transfer of ascites containing the 6-19 mAb into BALB/c mice induces, within 18 h, remarkable pathology characterized by skin vasculitis and acute glomerulonephritis associated with cryoglobulinemia. In order to evaluate the possibility of modulating the development of tissue lesions by an anti-Id antibody, we have raised an IgG2b anti-Id mAb specific to the 6-19 mAb. The cryoprecipitation of 6-19 mAb was completely inhibited in the presence of excess amounts of anti-Id mAb in vitro. In vivo, pretreatment of BALB/c mice with anti-6-19 anti-Id mAb inhibited development of skin vasculitis and glomerulonephritis induced by the 6-19 mAb. The cryoglobulin formation was markedly diminished due to enhanced elimination of the 6-19 mAb from the circulation. In contrast, pretreatment with an IgM anti-IgG3 rheumatoid factor mAb neither protected nor aggravated the development of tissue lesions. These results suggest possible implications in the anti-Id treatment of similar vascular diseases in man.