M-Dot-ELISA检测麻风家内接触者血清中麻风杆菌特异性抗原

用改良酶联免疫斑点１５试验（简称 Ｍ－Ｄｏｔ－ＥＬＩＳＡ）对麻风家内接触者（ＨＣ）血清进行了麻风杆菌特异性酚酚糖脂Ｉ（ＰＧＬ－Ｉ）抗原检测。７５例ＨＣ均经麻风杆菌明胶微粒凝集试验（ＭＬＰＡ）和酶联免疫吸附试验（ＥＬＩＳＡ）证实为麻风杆菌特异性抗体阳性者。结果表明：①７５例ＨＣ中抗原阳性者１６例，阳性率为２１％。接触多菌型麻风（ＭＢ）者的抗原阳性率（２８％）明显高于接触少菌型麻风（ＰＢ）者（０％），两者之间有极显著性差异（ Ｐ ＜０．０１ ）。②抗体弱、强阳性组的抗原阳性率和抗原量之间亦有极显著差异（Ｐ＜０．０１）。③４０例正常人及１０例经ＭＬＰＡ和ＥＬＩＳＡ检测证实特异性抗体阳性的非麻风患者血清标本，抗原检测均阴性。     

麻风杆菌重组α抗原的应用研究:α2-ELISA在麻风复发监测中的应用初探

目的：探讨以重组麻风杆菌α２融合蛋白为抗原的酶联免疫吸附试验（α２－ＥＬＩＳＡ）在监测麻风复发中的应用价值。方法：用已建立的α２－ＥＬＩＳＡ检测３１２例用ＤＤＳ单疗的麻风病治愈者血清中的抗α２融合蛋白抗体。结果：多菌型组α２抗体阳性率高于少菌型组,且在多菌型治愈者中的α２抗体滴度显示随着ＤＤＳ疗程的增加而降低的趋势。结论：提示α２－ＥＬＩＳＡ在多菌型麻风治愈者中可望成为一个麻风复发监测的血清学手段之一,且在麻风疗效的评价中有一定的意义。     

麻风菌重组融合蛋白α-抗原的血清学活性

用麻风患者的血清（Ｌ３８份、Ｂ１９份、Ｔ３６份）比较了α１－ＥＬＩＳＡ、α２－－ＥＬＩＳＡ与ＰＧＬ－Ｉ－ＥＬＩＳＡ法，结果各型患者血清中的抗体滴度分别为：α１－抗原＞α２－抗原＞ＰＧＬ－Ｉ，尽管Ｌ型的血清中抗α１－抗原的滴度明显高于其它两种抗原，但其阳性率在三种方法之间统计学上没有差别，分别为９７％，９２％和９４％；Ｂ和Ｔ型血清α１－ＥＬＩＳＡ法的阳性率明显高于其它两种方法，分别为８４％、５８％、６８％和４７％、３８％、３０％，以α１－ＥＬＩＳＡ法的敏感性最高。提示α１－ＥＬＩＳＡ在麻风血清诊断中有很大潜力，值得进一步扩大样本进行评价     

酶联免疫吸附试验检测麻风特异性抗PGL－1抗体

用麻风菌特异性酚糖脂－１（ＰＧＬ－１）抗原（ＮＴ－Ｐ－ＢＳＡ）作酶联免疫吸附试验（ＥＬＩＳＡ），分别检测麻风病人５０例（ＬＬ４０例。阳性１６例，占４０．０ｆ％；ＢＬ９例，阳性４例，占４４．４％；Ⅰ１例为阴性），总阳性２０例，占４０．０％。治愈老残病人１３３例阳性１９例，占１４．３％；家内接触者１０２例，阳性９例，占８．８％。并作了结核病人８７例及正常人群１７４例。结果表明ＮＴ－Ｐ－ＢＳＡ－ＥＬＩＳＡ法具有较高的敏感性和特异性，将有助于麻风的早期诊断、判愈、复发予测及亚临床感染和免疫流行病学等研究。     

酶联免疫吸附试验检测麻风特异性抗PGL－1抗体

用麻风菌特异性酚糖脂－１（ＰＧＬ－１）抗原（ＮＴ－Ｐ－ＢＳＡ）作酶联免疫吸附试验（ＥＬＩＳＡ），分别检测麻风病人５０例（ＬＬ４０例。阳性１６例，占４０．０ｆ％；ＢＬ９例，阳性４例，占４４．４％；Ⅰ１例为阴性），总阳性２０例，占４０．０％。治愈老残病人１３３例阳性１９例，占１４．３％；家内接触者１０２例，阳性９例，占８．８％。并作了结核病人８７例及正常人群１７４例。结果表明ＮＴ－Ｐ－ＢＳＡ－ＥＬＩＳＡ法具有较高的敏感性和特异性，将有助于麻风的早期诊断、判愈、复发予测及亚临床感染和免疫流行病学等研究。     

SACT的建立及其与PGL—I—ELISA的比较研究

用抗麻风菌特异酚糖脂Ｉ（ＰＧＬ－Ｉ）单克隆抗体Ｂ８Ｆ１ＩＶ建立了间接血清抗体竞争试验（ｓｃｒｕｍ ａｎｔｉｂｏｄｙ ｃｏｍｐｅｔｉｔｉｏｎ ｔｅｓｔ，ＳＡＣＴ），其性为９９％特异性为９６．５％。以本法与标准化的Ｃ ＰＧＬ－抗原的间接酶联免疫吸附试验（ＰＧＬ－Ｉ－ＥＬＩＳＡ）作了比较。所用麻风血清为１００例（ＬＬ２０、ＢＩ２０，ＢＢ２０，ＢＴ２０），正常人血清为２８例，结果显示丙地在检测ＭＢ病人上无     

用ND—O—BSA和PGL—I作抗原的家庭接解者麻风血清流行病学研究

作者于１９９１年４－５月对麻风高流行区广南县和文山县的麻风家庭接触（ＨＣＰ）７２３人，健康人（ＥＨＰ）１６３２人和甘肃省非麻风流行区健康人（ＮＨＰ）１３１人，用检测血清抗ＮＤ－Ｏ－ＢＳＡ和ＰＧＬ－１抗体水平的ＥＬＩＳＡ进行了血清流行病学研究，血清阳性标准是根据ＥＨＰ和ＮＨＰ的血清抗体水平，即ＥＨＰ的阳性标准分别为０．２３和０．２２５，ＮＨＰ的分别为０．１４和０．１７，根据ＮＨＰ的阳性标准（ＮＨＰＣ     

麻风菌重组融合蛋白α—抗原的血清学活性

用麻风２的血清（Ｌ３８份、Ｂ１９份、Ｔ３６份）比较了α１－ＥＬＩＳＡ、α２－－ＥＬＩＳＡ与ＰＧＬ－Ｉ－ＥＬＩＳＡ法，结果各型患者血清中的抗何不工分别为：α１－抗原〉α２－抗原〉ＰＧＬ－Ｉ，尽管ＬＧ 血清中α１－怕的滴度明显高于其它两种抗原，但其阳性率在三种方法之间统计学上没有差别，分别为９７％，９２％和９４％；Ｂ和Ｔ型血清α１－ＥＬＩＳＡ法的阳性率明显高于其它两种方法，分别为８４％、５８％、６８％     

麻风复发的免疫学监测及预防

对１５１５例麻风治愈者以酶联免疫吸附试验（ＥＬＩＳＡ），用全血滤纸干渍斑，连续５年检测其麻风菌特异抗体酚糖脂－免疫球蛋白Ｍ（ＰＧＬ－ＩｇＭ）。对第一年检测阳性者（２８１例）随机分成Ａ、Ｂ两组：Ａ组（１４０例）每年仅进行一次临床、细菌和免疫学监测；Ｂ组（１４１例）给ＭＤＴ一年，以后继续监测；阴性者为Ｃ组（１２３４例），每年一次临床、细菌和免疫学监测。第一年检测发现随着型别ＴＴ→ＬＬ，阳性率逐渐升高；     

麻风杆菌重组α抗原的应用研究:α2-ELISA的建立

目的;以重组麻风杆菌α２融合蛋白为抗原,建立酶联免疫吸附试验,即α２－ＥＬＩＳＡ。方法：用麻风病人和健康正常人的血清评价了α２－ＥＬＩＳＡ中有关试剂及最适浓度,其敏感性与ＰＧＬ－１－ＥＬＩＳＡ进行了比较。结果：（１）挥发性缓冲液醋酸铵碳酸盐为α２抗原最佳包被液;（２）重组α２融合蛋白的最适包被浓度为５μｇ／ｍｌ;（３）显色剂ＯＰＤ的敏感性大于ＴＭＢ;（４）ＹＳＳ ＭＱ ＵＧＭ Ｗ ＴＬＤ ＩＧＥ     

Evaluation of four semi-synthetic Mycobacterium leprae antigens with sera from healthy populations in endemic and non-endemic areas.

In order to determine the frequency of occurrence of antibodies to semisynthetic antigens of Mycobacterium leprae in clinically healthy nonpatient populations and to establish a 'baseline' for comparison with antibody frequencies in both patients with a history of leprosy and their contacts, ELISAs were conducted using representative sera from two areas: a leprosy endemic area, Cebu City, Philippines and a nonendemic area for leprosy Chicago, Illinois, USA. These sera were tested, by an indirect IgM ELISA, for the presence of antibodies reacting with four semisynthetic antigens based on the phenolic glycolipid I antigen of M. leprae: ND-O-BSA (natural disaccharide with octyl linkage to bovine serum albumin), NT-O-BSA (natural trisaccharide with octyl linkage to BSA), ND-P-BSA (natural disaccharide with phenolic ring linkage to BSA) and NT-P-BSA (natural trisaccharide with phenolic ring linkage to BSA). Using an OD reading > or = 0.16 as positive, the antigen with the lowest background seroreactivity was ND-O-BSA, which reacted with 5/398 (1.3%) sera from Cebu, and 3/426 (0.7%) sera from Chicago. A total of 10 (2.5%) of 398 sera from the endemic area reacted with at least one antigen and 5 (1.3%) sera reacted with all four semisynthetic antigens. Of the 426 sera from Chicago, 12 (2.8%) were reactive with at least one antigen and 3 (0.7%) were reactive with all four semisynthetic antigens. Mean ELISA values for the 22 positive sera for each antigen ranged from 0.17 to 0.3 OD units, while the mean values for all sera in each area ranged from 0.01 to 0.04 OD units for all four antigens. Reactivity of 14 of the positive sera to some antigens, but not all four semisynthetic antigens, indicated that the carrier and linker arms might be associated with this background reactivity. Investigation of alternative linker arms and carriers is warranted. We conclude that nonspecific background reactivity to the semisynthetic antigens representing the PG-I molecule of M. leprae is 0.7-1.3%, based on a > or = 0.16 OD cutoff value. From these data it was concluded that reactivity in individuals free of leprosy was low enough to warrant use of these antigens in a diagnostic setting, such as screening household contacts and highly endemic populations. When incidence and prevalence of leprosy are low, testing with these antigens would not be cost effective, unless applied to high risk individuals.(ABSTRACT TRUNCATED AT 400 WORDS)     

Detection of a Mycobacterium leprae cell wall antigen in the urine of untreated and treated patients.

A total of 90 leprosy patients, 12 household contacts and 10 normal subjects were studied for the detection of Mycobacterium leprae cell wall antigen in urine using monoclonal antibody (ML30A2 IgG). In untreated multibacillary leprosy (BL-LL) the M. leprae cell wall antigen could be demonstrated in the urine of 14 (64%) patients by immunofluorescence (IF) and 22 (100%) by ELISA. In untreated paucibacillary leprosy (TT-BT), it could be demonstrated in 3 (11.5%) and in 13 (50%) patients by IF and ELISA methods respectively. All but 1 household contact (later confirmed to have BL leprosy) and all 10 normal subjects' urine was negative for M. leprae cell wall antigen by both methods. The same antigen was, however, demonstrated in urine of 50% paucibacillary patients who had received 6 months of treatment and in 68% multibacillary patients who had received 24 months of WHO recommended multidrug therapy. M. leprae cell wall antigen assays in urine will not be useful in the follow-up of leprosy patients on multidrug therapy.     

Characterization of circulating immune complexes in leprosy patients and their correlation with specific antibodies against Mycobacterium leprae

Circulating immune complex (CIC) levels and their antibody and antigenic composition were evaluated in patients with leprosy as well as in any individuals living with them; they were precipitated with 3.5% polyethylene glycol (PEG) and, after affinity chromatography isolation and purification, analysed by sodium dodecyl sulphate—polyacrylamide gel electrophoresis (SDS-PAGE) and immunoblot with monoclonal antibodies (Mabs). The presence of CICs was demonstrated throughout the clinical and immunopathological leprosy spectrum at levels related to bacterial load, and in leprosy patients they showed a positive correlation with specific anti-PGL and anti-65 kDa antibodies. The isolation and analysis, however, failed to identity any Mycobacterium leprae antigenic components; although two specific antibodies anti-PGL-1 and anti-65kDa were identified as possible CIC constituents and may be potentially useful in the follow-up of leprosy patients, especially to chirk bacterial load evolution, PG1.-1 being an authentic antigen of this mycobacterium. Also, the involvement of 65 kDa in CICs, being homologous with the human heat shock protein (HSP) 60 kDa family, suggests an autoimmune mechanism in leprosy pathogenesis, Furthermore, those results support the inclusion of CIC anti-body reactivity studies to enhance the sensitivity of serology.     

Evaluation of anti-cardiolipin antibody and its cross-reactivity in sera of patients with lepromatous leprosy

Summary Using a sensitive and modified solid-phase radioimmunoassay for detecting anti-cardiolipin antibodies, sera of 45 patients with lepromatous leprosy were examined. Nine of the 45 (20%) showed positive levels of anti-cardiolipin antibodies. Inhibition tests revealed that these antibodies significantly cross-reacted with double-stranded (ds) DNA, but not with single-stranded (ss) DNA or extractable nuclear antigens (ENA). We describe the unique pattern of antibody cross-reactivity with cardiolipin and dsDNA in sera of patients with lepromatous leprosy.     

Use of non-conventional antigen: M. habana in detecting M. leprae antibodies from leprosy patients and contacts in FLA-ABS test

An indirect immunofluorescent (FLA-ABS) test has been developed to detect M. leprae specific antibodies in the active and subclinical cases of leprosy. An antigenically related mycobacterium, M. habana, was used as an antigen to detect M. leprae specific antibodies in the sera samples of leprosy patients. A comparison was made with M. leprae antigen using same set of sera samples. M. habana is capable of detecting anti-M. leprae antibodies in the serum samples of leprosy patients, previously absorbed with various mycobacterial antigens, cardiolipin and lecithin, almost to the same percentage as M. leprae. Possible use of M. habana antigen as an alternative to M. leprae, in the serodiagnosis of leprosy, has been discussed.     

Cyclosporine A treatment of leprosy patients with chronic neuritis is associated with pain control and reduction in antibodies against nerve growth factor

OBJECTIVES: Chronic neuritis (CN) is still a major problem in leprosy and is difficult to manage in patients who do not respond well to prednisone. In this study we (i) evaluate the efficacy of cyclosporine A (CyA) in controlling CN patients, and (ii) analyse the presence of anti-NGF antibodies in the sera of leprosy patients, and their behaviour during CyA treatment. DESIGN: This was an open, prospective, non-comparative study. Sixty-seven leprosy patients in three different institutions in Pará, Brazil were studied from January, 2001 to January, 2004. Of these, 47 had no CN and 20 were leprosy patients suffering from CN and taking at least 40 mg/day prednisone to control nerve impairment and pain. Patients received 12 months reducing course CyA starting at 5 mg/kg per day. The outcome measure was sensory impairment, assessed using Semmes-Weinstein monofilament examination (SWME), muscular force and spontaneous or palpation-related pain. RESULTS: Antibodies against NGF were detected in the sera of leprosy patients, which may explain the depletion of NGF in leprosy contributing to neuritis, inflammation and loss of cutaneous nociception. The levels of these antibodies in CN patients were slightly lower than in patients with no CN. However, anti-NGF titres in CN patients treated with CyA were lowered to levels similar to those in the normal subjects. There was also improvement in sensory impairment, muscular force and pain. CONCLUSIONS: These data suggest that anti-NGF antibodies are present in the sera of leprosy patients and may influence the outcome of neuritis, and that CyA might be a useful drug in controlling nerve impairment and pain in leprosy patients.     

A dot enzyme immunoassay for detection of IgM antibodies against phenolic glycolipid-I in sera from leprosy patients

A visual dipstick dot enzyme immunoassay (EIA) for diagnosis of leprosy is described. The assay is based on detection of IgM antibodies against phenolic glycolipid (PGL-I) in sera from leprosy patients. The antigen (PGL-I or synthetic disaccharide of PGL-I) was dotted on a nitrocellulose pad stuck on a plastic strip (dipstick). Sera were used at a dilution of 1:200. Peroxidase coupled mouse anti-human IgM monoclonal antibodies were used as the conjugate. A positive test gave a blue dot against a white background. The test was highly specific for leprosy, and was quite sensitive for detection of bacilliferous (BL/LL) leprosy. The antigen dotted and preblocked dipsticks stored at room temperature upto 4 months of observation period, were unable in the assay.     

Evaluation of Phenolic Glycolipid-I (PGL-I) Antibody as a Multidrug Therapy (MDT) Monitor

Since phenolic glycolipid-I (PGL-I) is an unequivocal marker for Mycobacterium leprae, this antigen has been a good candidate for the serodiagnosis and monitoring of the effectiveness of leprosy chemotherapy. The present study, a continuation of an earlier report, was undertaken to estimate PGL-I antibody titers in 40 leprosy patients 3 and 6 months after starting MDT. All the leprosy groups showed significant declines in anti PGL-I reactivity after 6 months. There was a good correlation between bacteriological indices (BI) and anti PGL-I antibody levels. Thus, PGL-I based serology may be useful in monitoring the response to multidrug therapy.