食蟹猴疟原虫和恶性疟原虫两种抗原检测不同疟区人群中间接荧光抗体的实用性

[目的 ]比较食蟹猴疟原虫 (P c.)和恶性疟原虫 (P f.)两种抗原在不同疟区人群疟疾抗体检测的实用性。 [方法 ]1997年 5～ 10月在海南省间日疟和恶性疟混合流行区及河南省单纯间日疟区用P c 和P f 两种抗原测试人群疟疾抗体。 [结果 ]在海南省间日疟和恶性疟混合流行区P c 和P f 两种抗原检测人群疟疾间接荧光抗体阳性率分别为 37 4%和 31 3% ,其阳性符合率为 83 9% ;河南省单纯间日疟流行区的P f和P c两种抗原阳性率分别为 2 3 0 %和 9 7% ,阳性GMRT分别为 42 9%和 2 9 3%。 [结论 ]间日疟和恶性疟混合流行区P f和P c两种抗原均可用于人群疟疾抗体的检测 ,而单纯间日疟地区则以P c 抗原为优。     

金标免疫渗滤法诊断试剂盒诊断血吸虫病的价值研究

目的　评价金标免疫渗滤法诊断试剂盒（DIGFA-kit）用于血吸虫病血清流行病学调查的价值。方法　采用DIGFA与ELISA单盲平行检测各期血吸虫病病人血清、非流行区人群血清、疫区人群血清和血吸虫病已控制地区人群血清共2304份。结果　DIGFA检测血吸虫抗体的敏感性为96.8%，对非流行区健康人群的特异性为100%；该法检测疫区居民血清508份，抗体阳性检出率为35.6%，在Kato法粪检阳性的81人中，其中抗体阳性79人，阳性符合率为97.5%；检测血吸虫病已控制地区人群血清，抗体阳性率为4.6%；根据DIGFA的敏感性和对非流行区人群的特异性，计算其Youdens指数为0.926。上述各项结果与ELISA比较，差异无显著性（P>0.05）。结论　DIGFA与ELISA有相似的敏感性和特异性，且方法简便、快速，胶体金标记物制备容易，试剂稳定，运送方便，在现场和临床诊断更具有优越性。     

用单克隆抗体——酶联免疫吸附试验双抗体夹心法检测人体疟原虫抗原的研究

采用兔抗恶性疟原虫(Fcc株)抗体包被,以现症疟疾病人滤纸血为检测样本,用两种单克隆抗体M_26-32。3F_9混合进行反应的ELISA双抗体夹心法检测红内期疟原虫抗原,取得较满意结果: 64份镜检阳性疟疾样本(海南岛恶性疟31份;江苏恶性疟16份,间日疟17份)和25份非疟疾对照样本测定结果,阳性和对照样本的平均增强率分别为77.5和     

快速诊断疟疾胶体金免疫层析试条方法的建立与评价

目的 建立一种能区分恶性疟的快速、简便诊断疟疾的胶体金免疫层析试条方法，并对其进行评价。 方法 筛选基于恶性疟原虫乳酸脱氢酶制备的单克隆抗体对，采用柠檬酸三钠还原法制备胶体金颗粒，标记筛选到的单克隆抗体F4H12、G4C9和D8F7，并将其吸附于样品垫；将单克隆抗体B2G10（针对恶性疟原虫与间日疟原虫）和D6A7（只针对恶性疟原虫）分别划线包被于同一硝酸纤维素膜适当位置，制成免疫层析检测试条。用该试条检测疫区非疟疾发热病人血样（107份）和内脏利什曼病患者血样（17份）以评价其特异性，检测确诊的疟疾患者血样（间日疟110份， 恶性疟54份）以评价其敏感性。均用单盲法检测。 结果 检测107份疫区非疟疾发热病人血样和17份内脏利什曼病患者血样，有119份显示为阴性，特异性约为96.0%；其中17份内脏利什曼病患者血样全部为阴性。检测164份疟疾患者血样，阳性153份，敏感性为93.3%，其中间日疟检出率为92.7%（102/110），恶性疟检出率为94.4%（51/54）。 结论 研制出的快速诊断疟疾胶体金免疫层析试条敏感性、特异性均较高。     

快速免疫色谱测试法诊断恶性疟的初步观察

目的：评价快速免疫色谱测试法（ＩＣＴ）在我国疟区门诊疟疾的适用性。方法：以镜检结果为对照，用ＩＣＴ方法检测门诊“四热”病人中的恶性疟。结果：ＩＣＴ检测恶性疟的敏感性和特异性分别为９４．７％和９０．３％，与间日疟无交叉，检测不同性别和民族人群阳性率间的差别无显著意义。结论：ＩＣＴ较镜检诊断恶性疟更为快速且简便，更适于在疟区门诊应用。     

快速免疫色谱测试法诊断恶性疟的初步观察

目的：评价快速免疫色谱测试法（ＩＣＴ）在我国疟区门诊疟疾的适用性。方法：以镜检结果为对照，用ＩＣＴ方法检测门诊“四热”病人中的恶性疟。结果：ＩＣＴ检测恶性疟的敏感性和特异性分别为９４．７％和９０．３％，与间日疟无交叉，检测不同性别和民族人群阳性率间的差别无显著意义。结论：ＩＣＴ较镜检诊断恶性疟更为快速且简便，更适于在疟区门诊应用。     

急性无并发症疟疾病人和脑型疟病人的抗子孢子抗体

作者于1980年7～12月间采自急性无并发症疟疾和脑型疟的病人血清各95份,作间接荧光试验,检测病人血清中抗恶性疟和抗间日疟的子孢子抗体IgG和IgM,抗体滴度>1:8为阳性。其中95份脑型疟病人血清     

套式/多重PCR方法应用于疟疾诊断与监测的初步评价

目的 与镜检法比较评价标签引物-套式/多重PCR（UT-PCR）在疟疾监测中的应用价值。 方法 在海南、云南省恶性疟和间日疟混合流行区和广西疟疾控制区的疟疾监测中,采集初诊为疟疾或疑似疟疾的发热患者的血片与滤纸血样400份,在双盲条件下比较镜检法与UT-PCR的初检结果,对结果不一致的血片再次镜检复查,同时对其滤纸血样重复PCR 2～3次;评估UT-PCR与镜检法的敏感性和特异性。 结果 400例发热患者血样中,镜检法初检检出疟原虫阳性234例,其中恶性疟125例,间日疟109例;UT-PCR检出疟原虫阳性235例,其中恶性疟124例,间日疟109例;恶性疟和间日疟混合感染2例。两法初检结果一致的血样占92.5%（370/400）,其中阴性154例,阳性216例（间日疟117例,恶性疟99例）。复查25份初检结果不一致的血样,包括镜检阴性PCR阳性11例,镜检阳性PCR阴性10例,镜检为恶性疟PCR为间日疟3例,镜检为间日疟而PCR为混合感染1例,其中15份与UT-PCR的初检结果一致,7份“假阳性”原因不明,仅3份为PCR的假阴性。根据复查结果评估PCR的敏感性为99.6%,特异性为98.8%。 结论 采用更敏感的UT?鄄PCR疟疾诊断方法有助于解决疟疾诊断与鉴别诊断中的疑难问题,提高疟疾监测的质量和效率。     

微量快速检测血吸虫抗体的斑点免疫金渗滤法试剂盒研制

目的 研制一种适用于现场使用的血吸虫抗体快速检测试剂盒.方法 通过对原有的斑点免疫金渗滤法全面改进,建立新的快速检测血吸虫抗体试剂盒.采用双盲法,检测慢性血吸虫病人血清100人份,正常人、乙肝患者及其他寄生虫病感染血清140人份.以敏感性、特异性、Youden指数、Kappa值等指标评价试剂盒的检测效果.结果 试剂盒的敏感性为92%,特异性为95.08%,Youden指数为0.87,Kappa值为0.87,与华支睾吸虫感染者血清交叉反应率为5%.结论 斑点免疫金渗滤法试剂盒血清用量少,反应快,敏感性、特异性较高,适用于现场血吸虫抗体检测.     

多重PCR种特异性检测恶性疟原虫和间日疟原虫方法的建立

目的建立恶性疟原虫和间日疟原虫种特异性检测的多蕈PCR方法,用于疟疾的检测和诊断.方法根据疟原虫18S核糖体小亚基ssRNA的基因序列设计合成8对11条引物,通过对恶性疟、间日疟患者及健康对照者血样的DNA进行扩增,选择出敏感性和特异性最佳的引物用于建立多重PCR方法,并用梯度变化的方法分别对引物浓度、复性温度、延伸温度和循环次数等反应参数进行比较分析,优化PCR反应条件.利用优化后的多重PCR埘采自云南和上海的139份疟疾患者血样和32份非疟疾患者血样进行检测,以镜检方法为金标准,分析多重PCR方法检测患者血样的敏感性和特异性.结果从8对11条引物中优选出2对共3条引物用于建立多重PCR.利用这3条引物进行多重PCR,一次反应即可完成对恶性疟原虫和间日疟原虫的种特异性鉴定.对疟疾和非疟疾患者血样检测结果显示,该方法检测患者血样的敏感性为97.8%,特异性为100%.结论多重PCR方法敏感、特异、可进行批量检测,适用于对人群的疟疾监测和疑似疟疾病例的诊断,并能鉴定恶性疟原虫和间日疟原虫虫种.     

An immunocytochemical test for the diagnosis of antibodies to Plasmodium vivax

An immunocytochemical peroxidase test (ICPT) has been developed to allow serological measurement of the antibody response to Plasmodium vivax by light microscopy. Acetone fixed P. vivax erythrocyte stages were used as source of antigen. The immunofluorescent antibody test (IFAT) was used as reference test. In testing sera from individuals infected with P. vivax in southeastern Venezuela a high correlation (100%) was obtained between the ICPT and the IFAT. There were cross reactions with sera from patients with malaria by P. falciparum but not with those from patients with other parasitic diseases. Antibody titres as measured by the ICPT showed a positive correlation with past P. vivax malarial experiences.     

Evaluation of a Malaria Antibody ELISA and Its Value in Reducing Potential Wastage of Red Cell Donations from Blood Donors Exposed to Malaria, with a Note on a Case of Transfusion-Transmitted Malaria

Background and objectives: Blood donations are often wasted for lack of a satisfactory procedure to evaluate donors potentially exposed to malaria. Materials and methods: We evaluated a commercial ELISA for the detection of antibodies to malaria and compared it with an immunofluorescent antibody test (IFAT). Results: When 5,311 sera from routine non-exposed donors were tested, 24 (0.45%) were positive by the ELISA, using a Plasmodium falciparum antigen. Seventeen were subjected to confirmatory testing but none were positive by IFAT. Of 1,000 donors potentially exposed in endemic areas 15 (1.5%) were repeatably reactive by ELISA. 10 of these were tested by IFAT and 2 were positive. When 150 patients attending the Hospital for Tropical Diseases in London with acute malaria were tested, 73% of those infected with P. falciparum were repeatably reactive for malarial antibodies by ELISA and 56% with Plasmodium vivax. Of 88 stored clinical sera tested by both IFAT and ELISA 56 were positive by IFAT and of these 52 (93°/0) were positive by ELISA. Conclusion: The ELISA is sufficiently sensitive and specific to screen at-risk donors. Its use could safely retrieve 40,000 red cell units currently discarded each year in Great Britain.     

双抗体夹心斑点金免疫渗滤法检测血吸虫循环抗原的现场应用

目的 探讨双抗体夹心斑点金免疫渗滤法（S－DIGFA）在现场的应用价值。方法 在原已建立的以抗26kDaGST基因重组蛋白多克隆抗体为捕获抗体兼测示抗体的S－DIGFA的基础上，应用该法检测血吸虫病中度流行区和传播阻断地区人群的血清共1388份，并以双抗体夹心ELISA（S－ELISA）作对照。结果 用S－DIGFA和S－ELISA平行检测血吸虫病中度流行区人群血清300份，阳性检出率分别为15．7％和17．3％；两法检测血吸虫病传播阻断地区人群血清1088份，阳性检出率分别为3．9％和3．7％。结论 S－DIGFA检测循环抗原可在现场扩大和作血清流行病学调查。     

Antibodies against malaria sporozoites in patients with acute uncomplicated malaria and patients with cerebral malaria

Serum samples from 95 patients with acute uncomplicated falciparum malaria (AM) and 95 patients with cerebral malaria (CM) were tested by the indirect immunofluorescent assay (IFA) for IgG and IgM antibodies against Plasmodium falciparum and P. vivax sporozoites. Forty-six (48%) CM patients were positive for antibodies against P. falciparum sporozoites whereas only 23 (24%) were positive for antibodies against P. vivax sporozoites (P less than 0.002). A similar result was obtained in AM patients. However, CM patients had significantly lower mean IgG anti-sporozoite titer for P. falciparum than did AM patients (P less than 0.05), especially when only anti-sporozoite antibody-positive CM and AM patients were compared (P less than 0.0005), suggesting that CM patients had relatively less exposure and were probably less immune to malaria than were AM patients. The persistence of anti-sporozoite antibodies also was investigated in paired sera taken 63 days apart from 108 patients with acute falciparum malaria. There were significant decreases in the mean antibody titers in the follow-up sera during the period of stay in the malaria-free area. It was proposed that determination of anti-sporozoite antibody be made as a substitute for, or in addition to, anti-blood stage antibody for seroepidemiological study of malaria, especially in the monitoring of the success of the malaria control program.     

Two-site pan-species monoclonal antibody ELISA for detection of blood stage malaria antigen.

A two-site pan-species monoclonal antibody sandwich ELISA (MAb-MAb ELISA) was developed to detect both Plasmodium vivax and P. falciparum antigens in whole blood impregnated on filter paper. In this assay, the plates were coated with pan-species MAb 3F9 and another pan-species MAb M26-32 conjugated with alkaline phosphatase was used for detection of bound antigen. The sensitivity of this assay was 5, 10 and 10 parasites per 10(6) erythrocytes for cultured P. falciparum, patient-derived P. vivax and P. falciparum, respectively. The coincidence rates for this assay were 93% (92/99) with healthy individuals and 93% (42/45) with microscopically confirmed vivax malaria cases. After two weeks treatment, 77.7% (14/18) of vivax malaria were still positive by this assay but with diminished level of reactivities [corrected].     

Evaluation of a malarial antibody assay for use in the screening of blood and tissue products for clinical use

BACKGROUND AND OBJECTIVES: A new recombinant Plasmodium antigen enzyme immunoassay (EIA) for the detection of malarial antibodies was evaluated for the screening of 'malaria-risk' blood and tissue donations. MATERIALS AND METHODS: A total of 13,269 donor and patient samples were tested by both the EIA and the standard diagnostic antibody immunofluorescence test (IFAT). RESULTS: A total of 114/138 (82.6%) samples from patients with P. falciparum and 11/13 (84.6%) samples from patients with P. vivax tested positive. A total of 714/13,053 (5.47%) samples from donors identified as 'malaria risk', owing to residency or travel, were reactive in the EIA. CONCLUSIONS: The assay is more sensitive than a previously implemented malarial antibody EIA (73% in acute P. falciparum and 56% in acute P. vivax infections). The sensitivity of this new EIA is comparable to that of the IFAT, and the specificity is sufficient to screen 'malaria-risk' donors.     

Antibody detection ELISAS for malaria diagnosis.

Parasite extracts of Plasmodium falciparum and P. chabaudi and three synthetic peptides from the P. falciparum MSA2 merozoite antigen were tested for suitability as antigens in an antibody detection ELISA using sera from malaria patients in Brisbane. The P. chabaudi extract was superior to P. falciparum extract for detecting P. vivax cases, while for P. falciparum cases the two parasite extracts were equivalent. Single peptide antigens were generally less sensitive than parasite extracts; however, peptides G3 and G7 were more sensitive than parasite extracts in detecting first attacks of P. vivax. Examination of isotype specific responses demonstrated that this may be explained by higher IgG responses to these peptides in first than in subsequent P. vivax attacks. Because of the differing antibody specificities in primary and secondary P. falciparum and P. vivax cases, the best sensitivity was achieved by using the combined results of assays with three antigens: P. chabaudi, peptide G3 and peptide G7. The combined sensitivity was 77.1% for P. falciparum and 88.6% for P. vivax acute cases with 91.1% specificity.     

恶性疟原虫重组乳酸脱氢酶胶体金免疫层析法检测试纸条的研制

目的 用氯金酸标记抗恶性疟原虫乳酸脱氢酶（LDH）单克隆抗体（McAb），研制用于诊断恶性疟原虫和间日疟原虫感染的胶体金免疫层析（GICA）试纸。方法 采用柠檬酸盐还原法制备胶体金颗粒、标记抗恶性疟原虫重组乳酸脱氢酶单克隆抗体，制成免疫层析检测试纸条。用该试纸条检测已知疟疾病人及健康人血样，鉴定方法的敏感性和特异性。结果 检测重组LDH抗原的最小量为5ng／ml；对30例恶性疟病人血样及40例间日疟病人血样进行检测，敏感性分别为83．33％和57．50％，检测100例健康者血样特异性为98．00％。结论 初步建立了同时诊断恶性疟原虫和间日疟原虫感染的GICA检测法。     

Two-site sandwich ELISA for detection of Plasmodium vivax blood stage antigens using monoclonal and polyclonal antibodies.

Two systems of sandwich enzyme-linked immunosorbent assay (ELISA), a two-site monoclonal antibody sandwich ELISA MAb-MAb sandwich ELISA) and a two site polyclonal-monoclonal antibody sandwich ELISA (PAb-MAb sandwich ELISA) for the detection of Plasmodium vivax antigens were developed. The assays showed good correlation with the level of parasitemia when tested against serially diluted P. vivax parasites (r = 0.937, and 0.997 for MAb-MAb and PAb-MAb sandwich ELISA, respectively), with the ability to detect as few as 6.68 parasites/10(6) erythrocytes and 2.69 parasites/10(3) erythrocytes, respectively. The MAb-MAb sandwich ELISA was specific, since it was positive only with P. vivax-infected erythrocytes from vivax malaria patients and negative when erythrocytes from 34 healthy individuals and 30 falciparum malaria cases were tested. In contrast, cross-reaction was found in the PAb-MAb sandwich ELISA when the plates were coated with polyclonal IgG and tested against the serially diluted P. falciparum SO strain antigen prepared from in vitro cultures. Comparison between the two systems of two-site sandwich ELISA showed that the MAb-MAb sandwich ELISA was superior to the PAb-MAb sandwich ELISA: (1) it gave a higher sensitivity when tested with serially diluted P. vivax antigen preparations from vivax malaria patients; (2) it gave a higher specificity when tested with the SO strain of P. falciparum from in vitro cultures, (3) it gave a lower absorbance value when tested with erythrocytes from healthy individuals. All 281 cases of vivax malaria already proven by microscopic examination were positive by MAb-MAb sandwich ELISA.(ABSTRACT TRUNCATED AT 250 WORDS)