BA－ELISA快速检测鼻咽分泌物中合胞病毒抗原的研究

本文采用自制的ＲＳＶ免疫血清以及国产试剂，通过特异性、敏感度和重复性试验，建立了检测ＲＳＶ抗原的生物素－亲和素－酶联兔疫吸附试验（ＢＡ－ＥＬＩＳＡ）。该法比常规的ＥＬＩＳＡ法敏感４倍。用此法检测５８份肺炎和毛细支气管炎患儿鼻咽分泌物（ＮＰＳ）中的ＲＳＶ抗原，结果与组织培养（ＴＣ）分离ＲＳＶ结果比较：其特异性和敏感性分别为９１．２％和９５．８％，两法诊断符合率为９３．１％。结果表明本法不但敏感、特异，而且简便、快速、经济，在收到标本２４小时即可得到结果，是一种适应于基层推广的ＲＳＶ感染诊断方法。     

抗HBs的双抗原夹心BA-ELISA检测和应用

<正> 血清抗HB_s效价的测定是考核乙肝疫苗效果和判断人体有无抗乙型肝炎免疫力的重要指示。因此寻找一种较为理想的抗HB_s测定方法正在引人注目。现有PHA和ELISA等方法敏感性较差,阳性检出率低。近年来建立的ABC-ELISA(以下简称ABC法)用以检测人血清中抗HBs,阳性检出率有所提高。但该法使用非特异性第二抗体,使在特导性放价放大的同时,非特异性效应也随之增大,从而影响该法的使用效果。双抗原夹心BA-ELISA是以抗原固相化,以生物素化HBsAg和亲和素-辣根过氧化物酶结合物为指示。从而在克服单纯双抗原夹心法(以下简称夹心法)阳性检出率低的同     

用ELISA法快速检测病毒性脑炎患儿脑脊液中单纯疱疹病毒特异性抗原

用单纯疱疹病毒单克隆抗体（ＨＳＶ－ＭｃＡｂ）ＥＬＩＳＡ法检测了６２例病毒性脑炎患儿脑脊液中ＨＳＶ特异性抗原（ＨＳＶ－Ａｇ），２０例其他脑神经系统疾患者。３２例脑炎患儿同时取脑脊液和血清作ＨＳＶ－ＩｇＧ抗体水平测定比较。病毒性脑炎患儿的ＨＳＶ－Ａｇ阳性检出率为２０．６％，与血清／脑脊液的ＨＳＶ－ＩｇＧ测定比较，灵敏性８４．６％，特异性９４．７％。用ＥＬＩＳＡ法检测脑脊液中ＨＳＶ－Ａｇ是一种简便、快速、灵敏、特异的方法，对疱疹性脑炎具有早期诊断价值。     

应用单克隆抗体Dot—ELISA检测血吸虫循环抗原的初步研究

检测感染宿主体内的循环抗原和循环免疫复合物可以确诊寄生虫感染和考核疗效。国内已有多篇检测血吸虫循环抗原方法的报道,但应用单克隆抗体检测血吸虫循环抗原的研究尚未见诸国内文献。作者应用抗日本血吸虫排泄分泌抗原(S·ESA)单克隆抗体进行斑点酶联免疫吸附试验 (Dot-ELISA)检测感染宿主血清循环抗原,结果表明:本法具有较高的敏感性和特异性。 材 料 和 方 法     

应用MAC-ELISA方法检测呼吸道合胞病毒IgM抗体

建立酶联免疫吸附试验(MAC-ELISA)检测呼吸道合胞(RS)病毒特异性IgM抗体方法,并与中和试验(NT)进行对比。 取35例患毛细支气管炎婴幼儿的急性期及恢复期双份血清共70份标本进行研究。MAC-ELISA测得急性期血清中RS病毒特异性IgM抗体的阳性率为27/35(77.14％),而恢复期血清中和抗体呈≥4倍升高者为23/35(65.71％)。凡NT阳性病例,MAC-ELISA也呈阳性;凡MAC-ELISA阴性病例,NT也呈阴性。另4例NT阴性而MAC-ELISA呈阳性。两者阳性符合率为88.57％。 本研究也观察了IgM抗体与年龄的关系。在6个月龄以前,MAC-ELISA的阳性率远远高于NT。 实验说明采用MAC-ELISA检测RS病毒特异性IgM是可靠的快速诊断方法。     

应用ELISA方法检测抗呼吸道合胞病毒免疫核糖核酸免疫活性的研究

本文报导在小鼠模型系统中,应用ELISA方法对兔抗呼吸道合胞病毒(RSV)免疫核糖核酸(i-RNA)诱生血清特异性抗体及抗体生成的动态进行观察。实验结果表明:抗呼吸道台胞病毒免疫核糖核酸(RSV-i-RNA)在小鼠体内具有诱生血清特异性抗体能力。RSV-i-RNA常规致敏后24h抗体水平明显增高,72h达高峰,11天左右降至正常水平,RNase处理使RSV-i-RNA诱生血清特异性抗体能力明显减低,而DNase、Pronase对其则无明显影响。     

单克隆抗体Dot—ELISA半定量检测细胞表面HLA—DR抗原

用抗HLA-DR单克隆抗体HB55对Daudi、H18、健康人外周血淋巴细胞及K562细胞进行Dot-ELISA测定,检测细胞表面HLA-DR抗原。结果显示带该抗原的细胞,都有程度不同的显色;不带抗原的细胞,显色阴性。用岛津CS—9000薄层扫描仪画出显色点的色谱峰。峰的面积表示显色点大小,峰高表示显色深浅,它们的数值随样品细胞数增加而线性上升。经分析,发现采用峰高计算较采用面积计算重复性更好;而且加样量及细胞浓度对样品扩散影响不大。本技术操作简单。细胞用量少,与已报道的Dot-ELISA方法比较,具有用色谱扫描方法客观量化结果的独特之处。该技术可对细胞表面MHCⅡ类抗原及其他抗原表达及其动态变化作分析。     

用BA－ELISA斑点法检测痢疾杆菌免疫小鼠GALT中的特异性抗体分泌细胞（ASC）

改进的ＢＡ－ＥＬＩＳＰＯＴ法使免疫酶斑更为清晰，保存时间延长。用此法检测了痢疾杆菌福氏２ａ经口及腹腔免疫后，小鼠派伊尔氏（ＰＰ）淋巴结、肠系膜淋巴结（ＭＬＮ）及脾脏（ＳＰＬ）中特异性ＩｇＡ、ＩｇＧ、ＩｇＭ抗体分泌细胞（ＡｎｔｉｂｏｄｙＳｅｃｒｅｔｉｎｇｃｅｌｌ，ＡＳＣ）的动态变化，得到有规律的结果：两种免疫途径均能在ＰＰ及ＭＬＮ中诱导出３类特异ＡＳＣ的显著升高，但口服导致的升高其持续时间较腹腔途径为短。此外，腹腔途径还能诱导ＳＰＬ中３种ＡＳＣ升高。     

水痘-带状疱疹病毒抗原的ELISA定量检测方法的建立

目的:建立水痘-带状疱疹病毒(VZV)抗原的双抗体夹心ELISA定量检测方法,用于质控VZV灭活疫苗研发和生产中抗原含量.方法:以VZV中和单抗5F6C8为包被抗体、8H5D1为酶标抗体,构建定量检测VZV抗原的双抗体夹心ELISA方法,并对本方法的特异性、灵敏度、准确性、线性和稳定性等性能进行分析.结果:建立的双抗体夹心定量检测VZV抗原的ELISA方法,线性范围为0.4 μg～13 μg/ml,相关系数为R2=0.994,定量限度为0.4.μg/ml;变异系数CV＜ 15％、准确性回收率介于87.5％ ～ 111.6％之间,稳定性37℃6天的回收率＞80％.与VZV以外的相关病毒样本没有交叉反应.结论:构建的VZV抗原ELISA定量检测方法的各项性能符合定量检测需要,可用于VZV灭活疫苗的研发和生产过程的抗原含量检测.     

呼吸道合胞病毒G基因的扩增

用未纯化的呼吸道合胞病毒细胞混和物，采用改良胍－热酚法提取病毒ｍＲＮＡ，反转录合成ｃＤＮＡ，经ＰＣＲ扩增得到大小约９２０ｂｐ的产物。经ＰｓｔＩ酶切表明其中含有ＰｓｔＩ酶切应点，证实其产物确为ＲＳＶＧ基因。此法简便、特异、第三、快速、。由于ＲＳＶ极不稳定，ＲＮＡ甚易降解，此法值得推广应用。     

A MONOCLONAL ANTIBODY POOL FOR ROUTINE IMMUNOHISTOCHEMICAL DETECTION OF HUMAN RESPIRATORY SYNCYTIAL VIRUS ANTIGENS IN FORMALIN-FIXED,PARAFFIN-EMBEDDED TISSUE

Four monoclonal antibodies (MAbs) with specificities for epitopes on human respiratory syncytial virus (RSV) proteins preserved after formalin fixation and paraffin embedding were identified in fixed and embedded virus-infected HEp-2 cell pellets. The MAbs bound epitopes on the fusion protein, the nucleoprotein, the phosphoprotein, and the M2 protein of the virus. Following high-temperature antigen unmasking, immunohistochemical staining revealed RSV antigens in the lungs of five of seven children who died with confirmed RSV infection and in none of nine children who died for other reasons, with no evidence of RSV infection. Staining was cytoplasmic, granular, and confined to epithelial cells. Intense staining was seen at the apex of ciliated bronchial and bronchiolar epithelial cells in all five positive cases. In one case, of pneumonitis, infected pneumocytes were present in the alveoli and in several cases, CD68-positive, cytokeratin-negative alveolar macrophages stained for viral antigens. These antibodies may prove useful in studies of the pathogenesis of RSV infection. © 1997 John Wiley & Sons, Ltd.     

Ⅰ型变态反应与呼吸道合胞病毒毛细支气管炎喘鸣的关系

本文从RSV特异的IgE抗体(RSV-IgE),嗜硷粒细胞和组织胺三方面,探讨了RSV毛细支气管炎喘鸣的发病机理。实验发现,RSV毛细支气管炎患儿的鼻咽分泌液(NPS)中RSV-IgE和组织胺显著升高,外周血嗜硷粒细胞绝对数升高,对 RSV抗原敏感性高,脱颗粒阳性率高。初步证实 IgE介导Ⅰ型变态反应参与了 RSV毛细支气管炎的发病。同时发现,RSV-IgE在患儿体内持续存在,这可能是 RSV毛细支气管炎喘鸣患儿在病后数年中反复出现喘鸣的原因之一。     

CLINICAL EVALUATION OF THE MYCOBACTERIOPHAGE-BASED ASSAY IN RAPID DETECTION OF MYCOBACTERIUM TUBERCULOSIS IN RESPIRATORY SPECIMENS

Context: Search for a cost-effective, rapid and accurate test has renewed interest in mycobacteriophage as a tool in the diagnosis of tuberculosis (TB). There has been no reported data on the performance of phage assay in a high burden, low-resource setting like Kanpur city, India. Aims: To assess the sensitivity and specificity of the FASTPlaque TB™ kit ability to impact the bacillary load in the phage assay and its performance in the sputum smear sample negative cases. Materials and Methods: The study involved a cross-sectional blinded assessment of phage assay using the FASTPlaque TB™ kit on 68 suspected cases of pulmonary TB against sputum smear microscopy by Ziehl-Neilsen staining and culture by the LJ method. Results: The sensitivity, specificity and positive and negative predictive values of the phage assay were 90.7, 96, 97.5 and 85.7%, respectively. The assay was negative in all the five specimens growing mycobacteria other than TB. The sensitivity of the phage assay tended to decrease with the bacillary load. Of the smear-negative cases, three were false negative, and all of which were detected by the phage assay. Smear microscopy (three smears per patient) had a sensitivity and specificity of 93 and 64%, respectively. Conclusions: The phage assay has the potential clinical utility as a simple means of rapid and accurate detection of live Mycobacterium tuberculosis bacilli; however, its performance has been inconsistent across various studies, which highlights that the assay requires a high degree of quality control demanding infrastructure and its performance is vulnerable to common adversities observed in “out of research” practice settings like storage, transport and cross-contamination.     

DETECTION AND CROSS-REACTION OF H-Y ANTIGEN BY HAEMAGGLUTINATION

Haemagglutination has been developed for the detection of H-Y antigen and antibodies by using mouse and rat red blood cells (RBC). The method is simple, specific and sensitive. With this method, the H-Y antigen was demonstrated on the surface of RBC from mouse, rat, hamster, rabbit, cat, chicken, trout and man, as a cross-reactive surface component. RBC of the quail and the cat differed from the other species in their ability to absorb the H-Y antibodies, indicating unknown differences in the H-Y antigen of these species. The female trout was found to carry the H-Y antigen and thus is assumed to be the heterogametic sex of this species. Trout serum contains factor(s) with a potential for agglutinating mouse and rat RBC. Female mice (C57B1/6J) and rats (DA) were immunized with various male tissues according to different schedules. All H-Y antisera contained autoantibodies when tested by haemagglutination. The problems involving the detection of H-Y antigen and the effect of autoimmunity in H-Y antiserum are discussed.     

DETECTION OF A t-COMPLEX ANTIGEN BY SECONDARY CELL-MEDIATED LYMPHOCYTOTOXICITY

Because of the inconsistency in published results concerning the serological detection of cell surface antigens coded for by the t-complex, a cell-mediated lymphocytotoxicity (CML) assay, secondary CML, was used in a search for t-antigens. By sensitizing C3H. Ttf (C3H. Brachyury, tufted) with the congenic strain C3H. Ttf/t^w18 splenic cells, a response against lipopolysaccharide (LPS) stimulated splenic cells from C3H. Ttf/t^w18 mice is obtained. The locus coding for the antigen detected by this reaction lies to the left of tf on the murine seventeenth chromosome. The secondary CML response to this antigen is H-2 restricted and detects an antigen on all t-haplotypes tested: t^w18, t^w18tf, t¹², t⁶, t^h2tf, and t^w5.