酶联免疫吸附试验检测弓形虫体IgA抗体及其临床应用的初步研究

试用弓形虫膜抗原ELISA对先天性弓形虫感染的畸形围产儿和后天获得性弓形虫感染者进行了特导性IgA抗体检测。11例先天性弓形虫宫内感染畸形围产儿的15份标本,6份检出IgA抗体,7份同时检出IgA;IgM抗体,两份仅检出IgM抗体。8份弓形虫抗体阳性的孕妇血清皆检出较强的IgA抗体。后天获得性弓形虫近期感染者32份血清,IgA抗体阳性3份,IgA、IgM抗体阳性8份,IgM抗体阳性21份;30份慢性感染者血清未有测到IgA抗体。对照组30份抗体阴性的孕妇血清及其围产儿的脐血检测结果抗体皆为阴性。IgA抗体是弓形虫感染早期的一个重要标志物,与IgM、IgG抗体同时检测,对临床急性弓形虫感染的诊断和疗效观察有着极其重大意义。     

弓形虫膜抗原ELISA检测抗弓形虫IgA抗体的方法及其临 …

目的 建立弓形虫膜抗原ＥＬＩＳＡ检测抗弓形早ＩｇＡ抗体的方法，并评价其临角定ＥＬＩＳＡ担适反应条件，应用该法检测３６例先天性，３２例近期和３０例慢性弓形虫感染者及１５例连续观察的弓形虫病人治疗前后的血清ＩｇＡ抗体。结果 ３６例先天性，３２例例近期和３０例一感染血清的ＩｇＡ抗体检出数分别为２６例、２１例和０例。１５例连续观察对象有８例在治后２月，６例在治后４月，、例在治后５月，其ＩｇＡ抗体全部阴转。     

先天性出生缺陷儿童弓形虫感染血清学调查

目的探讨儿童先天性出生缺陷与弓形虫感染的关系。方法应用弓形虫间接血球凝集试验(IHA)和弓形虫膜抗原酶联免疫吸附试验(ELISA)检测300例先天性出生缺陷儿童血清弓形虫抗体,并以35例因后天获得性外伤或车祸致残儿童作为对照。结果300例先天性出生缺陷儿童弓形虫抗体阳性66例,阳性率为22.00%;显著高于后天获得性外伤或车祸致残儿童的弓形虫抗体阳性率8.57%(χ2=3.54,P<0.05)。同时在66例弓形虫抗体阳性的先天性出生缺陷儿童家庭中,其父母至少有一方弓形虫抗体阳性的家庭达38户,占弓形虫抗体阳性的先天性出生缺陷儿童家庭的57.58%。结论弓形虫感染是儿童出生缺陷的重要因素之一,对优生优育夫妇积极开展弓形虫感染的血清学监测,将对提高人口素质、控制先天性弓形虫感染的发生具有重要意义。     

弓形虫伴发巨细胞病毒感染32例孕妇妊娠结局的分析

为了探讨弓形虫伴发巨细胞病毒感染对胎儿的严重危害,我们对孕妇进行了血清学ELISA方法筛选检测了上述两种病原体的血清抗体及弓形虫的循环抗原(CAg),发现两种检测方法呈阳性反应的32例孕妇均引起妊娠不良结局。因此,对引产胎儿进行了剖尸病理检查,32例中就有21例找到弓形虫,并发现绝大多数胎儿有脑积水,无脑畸形、胸水、腹水及其他先天性畸形等。     

两种抗原检测弓形虫病IgM的效果

目的观察弓形虫P30重组抗原及可溶性抗原检测弓形虫病IgM的效果。方法建立斑点免疫金渗滤法（DIGFA）。采用2种抗原分别包被在两条硝酸纤维素膜（NC）上,与待检血清中的相应IgM抗体结合,通过金标记兔抗人IgM直接显色,以检测弓形虫病IgM。2种抗原的DIGFA分别检测弓形虫IgM抗体阳性、类风湿、支原体肺炎、血吸虫病人和正常人血清。结果两种抗原建立的DIGFA检测试剂共检测5种血清样本166份,其中弓形虫感染IgM抗体阳性病人血清38份,弓形虫P30DIGFA敏感性为92.1%（35/38）,可溶性抗原DIGFA敏感性为81.6%（31/38）,差异无统计学意义（P〉0.05）。正常人群对照血清63份,检测全部阴性,特异性为100%。检测类风湿病人血清30份、血吸虫病人血清25份及支原体肺炎病人血清10份,仅类风湿病人血清出现阳性交叉反应,其中弓形虫P30DIGFA阳性2例,可溶性抗原DIGFA阳性3例。结论弓形虫P30重组抗原检测弓形虫病人血清IgM的DIGFA试剂可用于临床早期或急性期弓形虫病的诊断。     

中枢神经系统疾病中弓形虫感染的检测研究

目的 了解中枢神经系统疾病尤其是脑囊中病患者中弓形虫感染情况,研究弓形虫抗体、抗原检测方法的效果和意义。方法 应用ELISA和IFAT抗体检测方法及单克隆抗体（4C10、1E9）夹心ELISA抗原检测方法对270例脑囊虫病、154例疑似脑囊虫病、88例原发性癫痫患者和144例健康人群血清进行弓形虫抗体、抗原检测和比较。结果 ELISA和IFAT检测各组人群抗弓形虫抗体阳性率分别为15．93％、13．64％、12．50％和6．25％、14．07％、13．64％、14．77％和5．56％。各 组弓形虫抗原阳性率分别为2．22％（6／270）、3．25％（5／154）、2．27％（2／88）、0．00％（0／144）。13例循环的阳性者,其抗体滴度为1：1600者1例,等于或高于1：3200者12例。结论 中枢神经系统疾病在脑囊中病患者中弓形虫感染率明显高于健康人群,提示部分脑囊虫病患者并发或继发了弓形虫感染,或脑弓形虫感染误诊为脑囊虫病或原发性癫痫病;与IFAT相比,ELISA检测抗弓形虫抗体较客观、易于定量;当患者血清中抗弓形虫抗体滴度等于或高于1：1600时,提示有弓形虫现症感染的可能性,应进一步进行弓形虫抗原、DNA或其他病原学检查;单克隆抗体（4C10、1E9）夹心ELISA可应用于弓形虫感染的诊断。     

80例眼部疾患者弓形虫抗原抗体检测结果报告

近年来，国内外有关弓形虫感染与眼疾关系的报道日益受到重视。为此，我们自1998年7月以来，对我院部分眼部疾病患者进行了弓形虫循环抗原CAg和特异性IgM、IgA、IgG抗体的检测。现将结果报告如下。1材料和方法1．1检测对象为我院眼科临床确诊患有眼疾的部分患者，共80例，其中男36例，女44例；年龄18岁至74岁；职业为干部15人，工人32人，农民18人，教师8人，个体工商者7例。1．2检测标本每例患者抽取全血3ml，及时分离血清，置－40℃冰箱保存备检。1．3弓形虫酶联免疫吸附试验（ELISA）试剂盒由华美生物技术研究所提供。1．4方法弓形…     

先天性畸形儿母婴弓形虫感染的血清学研究

本文对先天性畸形儿母婴弓形虫感染进行了血清学研究.82例先天性畸形儿弓形虫抗体阳性率为32.89%,抗原阳性率为60.61%.79例畸胎儿产妇弓形虫抗体阳性率为25.21%,抗原阳性率为40.63%.两者弓形虫抗体阳性率均显著高于正常人群.77例病理组织诊断阳性的畸胎儿弓形虫抗体阳性率为41.56%,抗原阳性率为60.38%;其产妇弓形虫抗体阳性率为25. 81%,抗原阳性率为41.37%.弓形虫抗体阳性的畸形儿,其母血清抗体阳性率高达62.50%,而弓形虫抗体阴性的畸形儿其母亲血清阳性率只有18.20%,两组间差异非常显著.提示:弓形虫感染可能是造成先天性畸形的重要病因之一;母婴垂直传播是弓形虫先天性感染的途径.     

斑点免疫金渗滤法检测弓形虫感染效果

目的建立一种简捷、准确的弓形虫感染检测方法。方法将弓形虫P30抗原包被在硝酸纤维素膜上,与待检血清中的相应抗体结合,通过金标记Protein-A蛋白直接显色。结果用该试剂检测5种血清样本共221份,其中弓形虫感染病人血清58份,敏感性为96.55%(56/58)。正常人群对照血清68份,检测结果全部为阴性,特异性为100%。检测25份类风湿病人血清、18份血吸虫病人血清及22份肺炎支原体病人血清,仅有1例类风湿病人血清出现阳性交叉反应。结论用弓形虫P30为抗原,以金标记Protein-A蛋白为探针显色系统的斑点免疫金渗滤法,对检测弓形虫病人血清抗体具有较高的敏感性和特异性,且操作简单、快速。     

从噬菌体肽库中筛选弓形虫抗原模拟表位初探

以弓形虫感染兔血清免疫球蛋白（Ig）作为靶分子，免疫筛选噬菌体随机7肽库和12肽库。经3轮筛选，随机挑取70个噬菌体克隆用ELISA检测其特异性，共获得43个阳性克隆。对23个ELISA反应较强的克隆进行测序，根据序列分析选择有代表性的克隆A7作为模拟表位抗原进行ELISA检测。对47份弓形虫感染兔血清进行ELISA检测，结果32份为阳性，敏感性为68.1%；155份健康人血清中弓形虫抗体呈阳性者10份，特异性为93.5%。从噬菌体随机肽库中能筛选到弓形虫抗原模拟表位有诊断弓形虫病的潜在价值。     

IgA antibodies for diagnosis of acute congenital and acquired toxoplasmosis

An ELISA for IgA toxoplasma antibodies was positive in 12 pregnant women who seroconverted during gestation. Positive IgA titers were also noted in 10 individuals with biopsy-proven toxoplasmic lymphadenitis; the highest titers were noted in the first months following onset of clinical signs. Toxoplasma IgA antibodies were also demonstrable in 8 of 9 infants/fetuses with congenital toxoplasma infection. In some, IgM antibodies could not be demonstrated. Among 20 patients with AIDS and biopsy-proven toxoplasmic encephalitis, only 1 had IgA antibodies. None of 20 individuals with chronic toxoplasma infection had demonstrable IgA antibodies. Demonstration of IgA toxoplasma antibodies should be useful for diagnosis of recently acquired infection and for diagnosis of the infection in the fetus and newborn.     

弓形虫伴发巨细胞病毒感染32例孕妇妊娠...

In an attempt to investigate the harmful effects of the infection of Toxoplasma and associated with cytomegalovirus on the fetus infection acquired transplacentally during their mothers' pregnancy sera from 1000 pregnant women were tested for antibodies to both pathogens and the circulating antigen of Toxoplasma by using ELISA. The test was positive (titre > or = 1:60400) in 32 of 1000 cases. Judging from the sero-positive results together with the case history, clinical manifestations and B-scan results, the 32 cases were advised to end their pregnancy. Necropsy and pathological examinations of the foetuses revealed stillborn foetus, hydrocephalus, acephalus, pleural effusion, ascites and other congenital malformations. Toxoplasma gondii were found in the tissues of 21 fetuses.     

Diagnosis of congenital toxoplasmosis in Scotland

During a period of 10 years (1976-1986) 853 cases of toxoplasmosis were identified serologically in the Toxoplasma Reference Laboratory in Scotland. Suspected cases of infection in pregnancy (29 cases) and of congenital infection (27 cases) were rare but ophthalmic disease was common (224 cases). In two of the congenital cases, maternal infection was identified during pregnancy. The other 25 babies were investigated because of suspected congenital infection. Toxoplasmosis was confirmed retrospectively in 11/25 babies and excluded in two of them. The other 12 cases remain unclassified. Diagnosis of infection in babies was difficult although the introduction of an IgM capture ELISA was of some help. IgM was detected by ELISA in 3/13 congenitally infected babies but by immunofluorescence tests in only one of these. In two antenatal studies, involving 16,000 women, an additional 32 women were identified whose babies might have benefited from treatment or the availability of fetoscopy for early detection of congenital infection.     

Epidemiological and clinicoimmunological monitoring of toxoplasmosis in the Omsk Region

The toxoplasmosis epidemiological surveillance system is being improved in the Omsk Region. The prevalence of toxoplasmosis has been analyzed among 31475 inhabitants from the Omsk Region over 15 years (from 1992 to 2006). There was a 1.7-2.5-fold increase in the rates of infection in the normal and abnormal pregnancy women. IgM was detected in 7% of the adult patients with suspected toxoplasmosis, 5.2% of the physiological pregnancy women, and 2.2 times more frequently (p < 0.01) in the pregnant women with threatening interruption. The study of avidity showed that in pregnant females, primary infection occurred in 4.2% of cases; the signs of subacute toxoplasmosis were noted in 6.09%. The IgA detection rate was 33%. IgM was found in 7.2% of children, 7.8% having low-avidity antibodies. Subacute toxoplasmosis was established in 10.4% of the patients with central nervous system and eye lesions or hepatitis. In 38.1% of cases, toxoplasmosis was concurrent with active cytomegalovirus infection. The authors show it expedient to determine IgA and its avidity and to detect Toxoplasma antigens, as well as antibodies to different Toxoplasma antigens.     

Detection of congenital toxoplasmosis in utero with a polymerase chain reaction on amniotic fluid

To establish a prenatal diagnosis of congenital toxoplasmosis the PCR test was done on amniotic fluids from 47 women suspected of primary Toxoplasma infection during pregnancy. Fragments of Toxoplasma B1 gene were found in 5 examined samples. Positive tests were confirmed by mouse inoculation or by serologic testing of newborns. It was concluded that the PCR performed on amniotic fluid should be recommended for the prenatal diagnosis of congenital toxoplasmosis, however, negative results of the test can not rule out congenital infection.     

Prevalence of congenital cytomegalovirus and Toxoplasma antibodies in Thailand

Prevalence of CMV and toxoplasmosis antibodies were determined among normal newborn infants, suspected congenital neonates and pregnant women. Seropositive rates of CMV and toxoplasmosis were similar. Most of the pregnant women had CMV antibodies. The in-house ELISA for detection of Toxoplasma antibodies was developed and compared with the commercial kit with sensitivity and specificity of 90.47% and 96.74% respectively.     

Determination of IgG, IgM and IgA antibody responses in human toxoplasmosis by enzyme-linked immunosorbent assay (ELISA)

A 4-layer modification of ELISA for the determination of toxoplasma antibodies is described. In 103 Finnish blood donors 27 had antibodies against Toxoplasma gondii. One donor had IgM antibodies and IgA antibodies were found in 9. In patients with acute toxoplasmosis a vigorous IgG antibody response resulted in high antibody levels soon after infection, declining gradually to mean adult levels in approximately 2 yr. IgM antibodies appeared during the earliest phases of infection and disappeared as early as in 1 or 2 months in some cases and in most cases by the 6th month after infection. An IgA antibody response was also always seen. It was slower than the IgM response and could therefore be used to improve the laboratory diagnosis especially in cases where IgM antibodies had already disappeared and no further increase in IgG antibodies could be detected.     

先天性弓形虫病的诊断与治疗研究

目的刚地弓行虫可以通过胎盘屏障而形成垂直传播,并可引起先天性弓形虫病,早期诊断难并常常会错过早期治疗的机会.本文对PCR和IHA两种诊断方法进行了比较,并采用了磺胺嘧啶和螺旋霉素进行了治疗.方法于1993- 1996年间,对296例新生儿静脉血样采用PCR和IHA方法进行了检测.结果PCR法优于IHA法(P＜0.05);以磺胺嘧啶和螺旋霉素治疗的17例患儿全部治愈,而且其生长发育均正常.结论 PCR方法显示出了特异性高、敏感性强并可应用于早期诊断,以预防新生儿死亡和后遗症的发生.各种不同表现的先天性弓形虫病是可以被发现的,而且采用磺胺嘧啶和螺旋霉素可以治愈.     

Protective immunity in the rat model of congenital toxoplasmosis and the potential of excreted-secreted antigens as vaccine components

Toxoplasma infection is a major cause of severe foetal pathology both in humans and in domestic animals, particularly sheep. We have previously reported the development of an experimental model to study congenital toxoplasmosis in the rat. Here we demonstrate that, as in humans, total protection against congenital toxoplasmosis can be achieved regardless of the strain of Toxoplasma gondii used to infect rats, or when initial and challenge infections were carried out with different strains. Chronic infection is associated with a highly specific immunity that involves both B-and T-cell responses beginning at day 10 postinfection. The antibody isotype analysis revealed that whereas immunoglobulin (Ig)G2b is the major elicited isotype, no IgG1 antibodies are detected. T cell proliferation was assayed using crude Toxoplasma extracts or excretory-secretory antigens (ESA). The analysis of T cell supernatants showed the specific secretion of both interleukin-2 and interferon-gamma by activated T cells. Immunization of rats before pregnancy with either crude Toxoplasma extracts or with ESA elicited a B cell response that included antibodies of the IgG1 isotype and conferred on the newborns high levels of protection. Preliminary experiments of immunization using two HPLC-purified ESA, GRA2 and GRA5, conferred, a significant protection although to a lesser extent. This experimental model represents an attractive model for the identification of future vaccine candidates against congenital toxoplasmosis.