旋毛虫肌幼虫特异性抗原的鉴定

目的 寻找诊断旋毛虫病的特异性抗原。方法 应用SDS—PAGE和Western blot对旋毛虫肌幼虫可溶性抗原中的蛋白组分进行研究。结果 旋毛虫肌幼虫可溶性抗原经SDS—PAGE后显示29条蛋白带，分子量范围在112—12kDa之间，其中65、43、42、31、30、20、17、16kDa为主带。112、110、108、102、97、95、65、63、58、55、53、49、45、43、42kDa蛋白组分与并殖吸虫感染的大鼠及患者血清、华支睾吸虫病、血吸虫病及囊尾蚴病患者血清均具有明显的交叉反应带；24—20kDa蛋白组分只与旋毛虫感染的大鼠、小鼠及患者血清反应，而不与其它寄生虫感染的动物或患者血清、正常大鼠和小鼠及正常人血清发生交叉反应。结论旋毛虫肌幼虫可溶性抗原中24—20kDa蛋白组分为旋毛虫肌幼虫的特异性抗原，可用于旋毛虫病的免疫学诊断及血清流行病学调杏。     

旋毛虫肌幼虫排泄分泌物中特异性诊断抗原的研究

目的　寻找旋毛虫肌幼虫排泄分泌(ES)物中的特异性诊断抗原。　方法　应用SDSPAGE和Western印迹对旋毛虫肌幼虫体外培养18、30h后的ES抗原中的蛋白组分进行研究。　结果　旋毛虫肌幼虫培养18、30h后得到的ES抗原组分大致相同,两种ES抗原中主要蛋白带的分子量为112、110、108、97、53、49、45、42、35、23和16kDa。18hES抗原中的102、97、95和53kDa以及30hES抗原中的53、49、45和43kDa均与并殖吸虫病、华支睾吸虫病、日本血吸虫病及囊尾蚴病患者血清发生明显的交叉反应。ES抗原中的23kDa蛋白组分只与旋毛虫感染的大鼠、小鼠及患者血清反应,而不与上述其它寄生虫感染者、正常大鼠和小鼠及正常人血清发生交叉反应。　结论　旋毛虫肌幼虫ES抗原中的23kDa蛋白组分为旋毛虫肌幼虫的特异性抗原,可用于旋毛虫病的血清学诊断及血清流行病学调查。     

Diagnostic values of IgG4 in human gnathostomiasis

The diagnostic values of immunoglobulin G subclass antibodies from patients with gnathostomiasis were assessed by immunoblot technique. Antigen was prepared from crude extracts of Gnathostoma spinigerum advanced third-stage larvae obtained from naturally infected eels. The sera were obtained from 14 parasite-confirmed gnathostomiasis cases, 63 patients with other helminthic infections and 13 healthy controls. Nine prominent IgG4 reactive bands appeared with molecular weights of 94, 51, 47, 43, 38, 24, 21, 20 and 15 kDa. The diagnostic sensitivity of each of the nine reactive bands ranged from 100% to 64.3% in 14 parasite-confirmed gnathostomiasis cases. All (100%) confirmed cases recognized the 21 kDa antigenic band, but not other helminthic infections or parasite-free control. Recognition of 21 kDa antigen in G. spinigerum advanced third-stage larvae crude extracts is the most specific diagnostic marker for human gnathostomiasis, with 100% sensitivity and specificity. The 20 and 24 kDa protein bands were additional diagnostic bands for confirming diagnosis of infection where the 21 kDa band was faint. No specific binding of IgG1, IgG2, or IgG3 antibodies was observed in any sera from confirmed gnathostomiasis cases.     

Antigenic profile,isolation and characterization of whole body extract of Paramphistomum gracile

An antigenic component of adult Paramphistomum gracile was characterized by means of indirect enzyme‐linked immunosorbent assay (indirect ELISA), sodium dodecyl sulphate–polyacrylamide gel electrophoresis (SDS‐PAGE) and immunoblotting using sera from cattle naturally infected with P. gracile, Eurytrema pancreaticum, Fasciola gigantica, Moniezia benedeni, strongylids, Trichuris sp. and Strongyloides sp. The whole body (WB) extracts of P. gracile were fractionated by gel filtration chromatography in a Sephadex G‐200 column. It was found that the WB extract fractions, F1–F3 were highly antigenic, F5 was moderately antigenic and F4 was poorly antigenic. For SDS‐PAGE and immunoblotting, the antigenic molecules of WB extract and all five fractions were mostly at molecular weights (MW) ranging from 12 to 150 kDa. One antigenic protein of 16 kDa detected in WB extract and F1–F3 was found to give a consistent reaction with sera from infected cattle. The antigenicity of the purified 16 kDa protein was confirmed by immunoblotting and indirect ELISA using a pool of sera and individual serum samples from infected cattle (at 1 : 78 125 dilution) and hyperimmunized rabbit (at 1 : 390 625 dilution). This finding suggests that the 16 kDa protein may be a potential antigen for the immunodiagnosis of cattle paramphistomosis caused by P. gracile.     

Studies on the 52 kDa antigen of Salmonella typhi: physicochemical stability, purification by affinity chromatography and immunochemical specificity.

The 52 kDa specific protein antigen of Salmonella typhi, as identified by monoclonal antibodies (Ekpo et al. 1990) has been studied with respect to its physicochemical stability, purification by affinity chromatography and immunochemical specificity. It was found that the 52 kDa protein was degraded into smaller antigenic fragments of MW 30-51 kDa when treated with acetone, ethanol, sodium thiocyanate, 0.3M sodium chloride and Veronal and Tris buffers. The exact chemical nature of the degradation of the protein under these conditions is not known but digestion by conventional proteases and dissociation of the non-covalent subunit type have been ruled out. It is proposed that the degradation may be the result of yet unidentified enzyme(s) which become activated by various physical or chemical treatments. Affinity chromatography using a specific monoclonal antibody has been carried out in an attempt to purify the 52 kDa protein. The binding of S. typhi protein to the column was saturable at 65.6 microgram protein/ml gel. The amount of S. typhi protein adsorbed on the column was 0.51% of the total sonicated cell protein. SDS-PAGE of the immunoadsorbent purified protein revealed bands at Mr 15-58 kDa, indicating that the protein obtained had been severely degraded. However, Western blot of the purified protein stained with a specific monoclonal antibody and with rabbit polyclonal antibody against S. typhi showed striking similarity, indicating that the protein obtained was close to immunochemical purity. The 52 kDa protein purified by affinity adsorbent was used as an antigen for the detection of specific IgM in sera of patients. It was shown that sera of patients infected with S. typhi as well as those infected with other bacteria, contained specific IgM against the 52 kDa protein. Thus, it appears that the 52 kDa protein contains species specific as well as cross-reacting epitopes. The possible development of specific diagnosis of S. typhi based on the present experimental results in discussed.     

弓形虫速殖子表膜蛋白的组分和抗原性分析

目的试验分析弓形虫速殖子表膜蛋白的组分及其抗原性。方法用聚丙烯酰胺凝胶电泳分析弓形虫速殖子表膜蛋白的组分和用弓形虫病人阳性血清及兔抗弓形虫表膜抗原血清识别硝酸纤维膜载体上的速殖子表膜抗原的免疫反应靶位。结果弓形虫速殖子表膜抗原的独特蛋白区带分子为16和17kDa,弓形虫病人特异性抗血清识别的速殖子表膜抗原的免疫反应位点为16kDa,免抗血清识别出64,35,32,26,16kDa等8个位点。结论弓形虫速殖子表膜蛋白存在特异的抗原决定簇和可被特异性抗体识别的受体,证明速殖子表膜蛋白具有较好的抗原性,此对于弓形虫病的免疫学诊断具有重要的意义。     

Immunoblot analysis of a 10 kDa antigen in cyst fluid of Taenia solium metacestodes

The diagnostic value of a 10 kDa subunit of 150 kDa protein in cyst fluid (CF) of Taenia solium metacestodes was evaluated. Immunoblot analysis revealed that most sera from patients with neurocysticercosis recognized the 10 kDa subunit strongly (209/217 cases, 84.6%), while a few sera from individuals with other parasitic diseases including alveolar echinococcosis (AE, 2/20, 10%) and cystic echinococcosis (CE, 2/25, 8%) showed faint reactions. Sera of cases with other parasitic diseases, especially AE and CE, exhibited cross reactions against other bands in CF. Both differential immunoblot and immunoprecipitation analyses showed that the 10 kDa subunit was the most specific to cysticercosis and highly antigenic, whereas other components at 20–40, 64, 95 and 106 kDa in CF were cross-reactive. IgG subclass ELISA and immunoblot demonstrated that both IgG4 and IgG1 reactions were predominant in neurocysticercosis and recognized the 10 kDa .     

旋毛虫、卫氏并殖吸虫及华支睾吸虫之间共同抗原的研究

目的鉴定旋毛虫、卫氏并殖吸虫及华支睾吸虫之间的共同抗原,避免血清学诊断这3种寄生虫病时的交叉反应。方法应用十二烷基硫酸钠-聚丙烯酸胺凝胶电泳(SDS-PAGE)和免疫印渍(Western blot)对旋毛虫肌幼虫、卫氏并殖吸虫成虫及华支睾吸虫成虫可溶性抗原中的蛋白组分进行研究。结果旋毛虫肌幼虫、卫氏并殖吸虫成虫及华文睾吸虫成虫3者相同蛋白带的分子量为108、65、53、43、42、31、25、16 kDa。免疫印渍结果表明,旋毛虫和卫氏并殖吸虫抗原中分子量为 65、58、53kDa的蛋白带均与旋毛虫感染的大鼠、小鼠及患者血清、肺吸虫感染的大鼠和患者血清发生反应;旋毛虫和华支睾吸虫抗原中分子量为108kDa的蛋白带均与旋毛虫感染的大鼠、小鼠及患者血清、肝吸虫病患者血清发生反应;而3种可溶性抗原中的53kDa与本实验中所有寄生虫感染的动物和患者均有交叉反应。结论 65、58、53 kDa蛋白为旋毛虫和卫氏并殖吸虫的共同抗原,108kDa蛋白为旋毛虫和华支睾吸虫的共同抗原,而53 kDa蛋白为这3种寄生虫的共同抗原。     

部分肠道线虫病的免疫交叉反应研究

以ＳＤＳ－ＰＡＧＥ酶联免疫印渍技术对蛔虫、钩虫、鞭虫及犬弓蛔虫的免疫交叉反应进行了研究。结果显示，钩虫病、鞭虫病及犬弓蛔虫病患者血清都与人、猪蛔虫可溶性成虫抗原发生较强的免疫反应。人蛔虫可溶性成虫抗原和猪蛔虫可溶性成虫抗原间无明显差别。人、猪蛔虫可溶性成虫抗原与钩虫病患者血清反应的主要抗原成份分子量为３５、４４、５２．５、６４和８５ｋＤａ。两种抗原与鞭虫病患者血清反应带位于２２．２ｋＤａ和２０５ｋＤａ之间。两种抗原与犬弓蛔虫病患者血清反应形成多条反应带，其抗原分子量在３５．５ｋＤａ和１９５ｋＤａ之间。本研究表明，在某些线虫之间的确存在免疫交叉反应。     

Antigenic recognition of rats infected with Gnathostoma hispidum: comparison of antigens derived from the advanced third stage larvae of Gnathostoma hispidum and Gnathostoma doloresi

To increase understanding of immune response of host against G. hispidum, antigenic recognition of rodent gnathostomiasis were identified by the immunoblot method after PAGE of the advanced third stage larvae of G. hispidum and G. doloresi. The molecular weight of the major antigenic bands of G. hispidum extracts were 35 kDa and 16 kDa, and of G. doloresi were 48 kDa, 37 kDa and 35 kDa. Of these bands, the 35 kDa antigen of G. hispidum and G. doloresi was recognized with the sera of rats infected with G. hispidum 5 weeks after inoculation. Lecting staining with endoglycosidase-H or hydrolytic degradation on nitrocellulose strips revealed that the 35 kDa band of G. hispidum possessed the mannose-rich oligosaccharides. On the other hand, the band of G. doloresi had the hybrid-type glycopeptides. The larval extracts and excretory-secretory materials of G. hispidum were not antigenitically identical.     

Evaluation of IHA, ELISA and Western Blot tests in diagnosis of pulmonary cystic hidatidosis

Pulmonary cystic hidatidosis caused by the larval stages of Echinococcus granulosus is a common parasitic disease in Turkey and throughout the world. In this study IHA, ELISA and Western Blot (WB) tests were performed with a panel of 59 sera from 31 surgically confirmed pulmonary cystic hidatidosis patients, 18 patients with pulmonary disease other than cystic hidatidosis and 10 healthy individual. The overall sensitivity of the IHA, ELISA and WB tests used for the serodiagnosis of pulmonary cystic hidatidosis were found as 96.7%, 87.1%, 100% and the specificities were 82.2%, 89.2% and %85.7, respectively. Using the WB test 8-12 kDa, 24 kDa and 124 kDa bands were detected as valuable for surgically confirmed patients' sera. One or more of these bands were also detected in sera of four patients with other pulmonary diseases false-positively. In conclusion conventional serologic test like IHA and ELISA is valuable in diagnosis of pulmonary cystic hidatidosis, also evaluation of some specific bands in WB would contribute to the diagnosis.     

棘球蚴感染绵羊并诱发休克期间特异性IgG、IgE抗体对特异性抗原的识别

目的 探讨细粒棘球蚴(E.g.)囊液粗制抗原和B抗原(EgB)识别绵羊感染E.g.后及诱发过敏性休克期间特异性IgG和IgE抗体的特异性反应,并对抗原特性及分子量进行描述。 方法 制备E.g.囊液粗制抗原及EgB,应用免疫印迹技术检测经剖杀证实感染E.g.并诱发过敏性休克的20只绵羊血清特异性IgG和IgE抗体对两种抗原的抗体反应性,并对抗原特性和分子量进行描述。 结果 特异性IgE抗体与耐热、低分子量的EgB(8、12和16 kDa)抗原未见明显的反应条带,而与E.g.囊液粗制抗原在43 kDa处可见明显的反应条带。IgG抗体与E.g.囊液粗制抗原未见明显的反应条带,但与EgB抗原反应后在31、43和66.2 kDa处可见较为明显的反应条带。 结论 EgB可能不是特异性IgE抗体的特异性抗原,而是IgG抗体的特异性抗原。E.g.粗制囊液抗原中含有特异性IgE抗体的特异性抗原组分,其分子量大于43 kDa,可能是导致棘球蚴病患者过敏性休克的主要致敏原。     

Characterization of Campylobacter cinaedi and C. fennelliae antigens and analysis of the human immune response

Whole-cell protein patterns of Campylobacter cinaedi and C. fennelliae, identified by SDS-PAGE, differed from one another and from five other catalase-positive Campylobacter species. When immunoblotted with rabbit antisera, C. cinaedi and C. fennelliae showed antigenic profiles that were highly species-specific and had little or no cross-reactivity with antigens of the other Campylobacter species tested. Antibody responses of 13 homosexual men with C. cinaedi infection demonstrated consistent reactions with five antigens (96-41 kDa). All human sera tested, including those of 10 culture-negative individuals, reacted with the C. cinaedi 59- and 64-kDa bands, but the overall responses of infected individuals were much stronger. The antigenic profiles of C. cinaedi and C. fennelliae can be used to distinguish these species from other Campylobacter species. At least six C. cinaedi antigens are immunogenic in humans.     

SDS－PAGE酶联免疫印渍实验对人、猪蛔虫抗原特性的分析

以ＳＤＳ－ＰＡＧＥ方法对人蛔虫和猪蛔虫成虫可溶性抗原分析发现，在分子量１４ｋＤ～２３０ｋＤ之间两种抗原有许多相同蛋白带，未见明显的特异性蛋白带存在。应用ＳＤＳ－ＰＡＧＥ酶联免疫印渍方法发现以该两种抗原制备的兔抗血清均能从两种抗原中辨认出多个蛋白带。猪蛔虫成虫可溶性抗原中分子量２１．７ｋＤ和３４ｋＤ蛋白带能被部分兔抗血清所识别，而人蛔虫成虫可溶性抗原未出现这２条反应带。以人蛔虫感染者血清进行酶联免疫印渍也发现人蛔虫、猪蛔虫成虫可溶性抗原相似。两抗原中许多蛋白带可被病人血清所识别，其主要蛋白带为６６ｋＤ，提示蛔虫病人血清中含有大量抗体。另外，其中一份病人血清能从猪蛔虫成虫可溶性抗原中识别出４８．５ｋＤ蛋白带，而在人蛔虫抗原中未发现有该蛋白带。     

日本血吸虫与华支睾吸虫和姜片虫之间的交叉抗原及其血清学反应性

应用ＳＤＳ－ＰＡＧＥ分离ＡＷＡ、ＣＳＡ和ＦＳＡ三种抗原的组分蛋白后，再以酶联免疫印迹技术（ＥＬＩＢ）进一步鉴定其组分蛋白的特异性蛋白带，分别显示１２条、１９条和２１条，其分子量（ＭＷ）范围分别为１３～６４ｋＤｅ、１６～１９８．５ｋＤａ和１４～１５０ｋＤａ。ＡＷＡ组分蛋白与兔抗ＳＥＡ免疫血清出现３条交叉反应带．ＭＷ为１７ｋＤａ、１４ｋＤａ和１３ｋＤａ，与抗ＣＳＡ的兔血清及华支睾吸虫病人血清显示３条（３８ｋＤａ、１７．５ｋＤａ及１７ｋＤｅ）交叉反应带．与抗ＦＳＡ的兔血清及姜片虫病人血清呈现５条（６４ｋＤａ、５９ｋＤｅ、５３ｋＤｅ、１７．５ｋＤａ和１７ｋＤａ）交叉反应带。实验结果表明，日本血吸虫成虫不仅与其虫卵之间存有交叉抗原，而且与华支睾吸虫和姜片虫成虫之间也存有交叉抗原及血清学交叉反应性。本研究为今后制备特异的血吸虫病血清学诊断抗原，提供了科学依据。     

日本血吸虫卵与卫氏并殖吸虫成虫可溶性抗原内的共同抗原组分研究

应用凝胶电泳和免疫印斑技术对日本血吸虫卵与卫氏并殖吸虫成虫的水溶性抗原(JSEA、PAA)和尿素溶性抗原(JEA-u、PAA-u)进行了抗原蛋白组分比较分析。结果显示:两者的特异性抗原显色主带,JSEA、JEA-u分别有8条和3条,PAA、PAA-u分别有9条和2条;其中JSEA、JEA-u分别有1条和2条,PAA、PAA-u分别有1条和2条对异种抗血清呈现交叉反应显色带。表明两种吸虫的可溶性抗原均存在共同抗原组分。慢性血吸虫病患者血清与并殖吸虫成虫抗原的交叉反应仅见于PAA,而未见于PAA-u,提示PAA-u在避免交叉反应和抗原纯化方面比PAA较为优越。     

犬蛔虫眼病病人血清中特异性IgG和IgE抗体水平的测定(英文)

目的研究犬蛔虫眼病病人血清中特异性IgG和IgE的抗体的水平。方法应用ELISA和Western-blotting。从东京医科齿科大学国际环境寄生虫学分野实验室储存的脉络膜炎的眼病病人血清中随意抽取105份血清,检测犬蛔虫抗体。结果通过ELISA法,105份血清中有82份IgG和IgE均为阴性(78 .1 %) ,12例血清仅IgG阳性(11 .4 %) ,3例血清仅IgE阳性(2 .9 %) ,8例血清IgG和IgE均为阳性。所有的IgG和IgE阳性的血清,进一步进行Western-blot。IgG的反应带分布在幼虫分泌排泄抗原分子重量(97 .2 -14 .3kDa)的整个范围内,IgE的反应带分布在幼虫分泌排泄抗原分子重量(97 .2—14 .3kDa)的相对狭窄的范围内(29 -45kDa)。结论在这个研究中,我们清楚的说明了一些脉络膜炎病例血清中IgE抗体比IgG抗体具有特异性,因此IgE在眼犬蛔虫病的诊断中具有特异性,IgE在眼犬蛔虫病诊断中的价值还要进行进一步的研究。     

Antibodies to somatic L3 antigen not protective against Brugia malayi infection

Western blot analysis of infective larvae (L3) antigen of Brugia malayi were performed on 200 sera from six groups of individuals: 36 samples from B. malayi microfilaremic individuals; 10 samples from individuals with elephantiasis; 50 and 20 samples from amicrofilaremic individuals in a B. malayi endemic area with no anti-filarial IgG4 antibodies (towards microfilaria and adult worm antigens) and samples with high titres of the anti-filarial IgG4 antibodies respectively; 50 samples from non-endemic normals and 34 samples from geohelminth-infected individuals. After protein transfer, PVDF membrane strips were successively incubated with blocking solution, human sera, monoclonal anti-human IgG4 antibody-HRP and developed with luminol chemiluminescence substrate. 28/36 (78%), 1/10 (10%) and 16/20(80%) of sera from individuals with microfilariae, elephantiasis and amicrofilaremic individuals with high titers of anti-filarial IgG4 antibodies respectively recognized L3 antigenic epitopes; the dominant and consistent antigenic bands were of approximately MW 43 kDa, 14 kDa, 15 kDa and 59 kDa. The rest of the sera were unreactive. This study showed that microfilaremics may or may not mount a notable antibody response to somatic L3 antigens, thus lending evidence that antibody response to this antigen is not protective against establishment of Brugia malayi infection.     

日本血吸虫各期虫体的酶联免疫电转移印迹分析及抗原定位研究

本文采用酶联免疫电转移印迹技术（EITB）分析、比较日本血吸虫不同发育时期虫体特定的组分蛋白分子、雌虫、雄虫和虫卵抗原分别与相应的雌、雄虫和虫卵免疫血清反应呈现17、20和8条蛋白带,这三种抗原与其它不同时期虫体免疫血清反应,三者间及彼此间均出现交叉反应,而雌、雄虫和虫卵抗原与其它寄生虫感染血清作用没发现有叉及反应。应用间接荧光抗体试验（IFA）和免疫酶染色试验（IES）对日本血吸虫抗原进行定位研究,其结果相似。血吸虫主要抗原物质来源于成虫表皮、肠上皮和卵内的毛蚴.     

细粒棘球蚴生发细胞特异性抗原的ELIB分析

应用SDS－PAGE和免疫印渍试验（EITB），证明体外培养的细粒棘球蚴生发细胞和囊液含有特异性抗原，分子量为52kDa和38kDa，而囊壁则含有此2种抗原。生发细胞的52kDa抗原可识别感染小鼠和包虫病患者血清的特异性抗体，对囊虫病人和正常人血清则无反应条带。