A Series of Serological Tests for the Detection of Antigens and Specific Antibodies in Deep-Seated Candidosis: Experimental Aspects/Eine Gruppe von Antigen- und Antikörpertests zur Serodiagnose von tieflokalisierten Candida-Mykosen: Experimentelle Aspekte

Summary: We describe a series of six serological tests for the diagnosis of deep-seated candidosis. The array comprises two commercial tests (antigen test, Ramco Inc., and antibody test, Roche), as well as four enzyme immunoassays which have been developed in this laboratory: an antigen test for detection of Candida-proteinase, the corresponding assays for monitoring of anti-proteinase antibodies, and two assays for monitoring of IgG and IgM against heterogenous metabolic antigens of C. albicans. The highly sensitive and specific proteinase antigen-test tolerates samples with high concentration of serum proteins. Proteinase antigen was detected in 10 out of 11 normal mice after intravenous infection with C. albicans blastospores. The proteinase antigen peaked between the second and fourth day after infection. A rise in corresponding antibodies was observed in all animals. No proteinase antigen was detected in sera of healthy human individuals; anti-proteinase antibody titers in these sera amounted up to 1:8000. In related ELISAs, using metabolic fungal antigens, titer values of specific IgG and IgM amounted to 5120 and 1280, respectively. The six tests were carried out in an comparative study under diagnostic conditions, the results of which are the subject of a forthcoming communication. Zusammenfassung: Ein Satz von sechs serologischen Tests für die Diagnostik der tiefen Candida-Mykosen wird vorgestellt. Die Gruppe schließt zwei kommerziell vertriebene Testbestecke ein (Latex-Agglutinationstest zum Antigennachweis, Ramco Inc., und Hämagglutinationstest zum Antikörpernachweis, Roche). Vier weitere Enzymimmuntests wurden von uns entwickelt: Ein Antigentest zum Nachweis von sekretorischer Candida-Protease, ein entsprechender Test zum Nachweis von Antikörpem gegen Candida-Protease, und zwei Assays zum Nachweis von IgG-bzw. IgM-Antikörpem gegen heterogene metabolische Antigene von C. albicans. Der empfindliche spezifische Protease-Antigentest toleriert hohe Konzentrationen unspezifischer Serumproteine und kann deshalb auf Serumproben in geringer Verdünnung (z. B. 1:20) angewandt werden. Protease-Antigen war in 200 fach verdünnten Seren von 10 aus 11 intravenös infizierten NWNI-Mäusen nachweisbar. Die höchste Antigen-Konzentration trat zwischen dem 2. und 4. Tag nach Infektion auf; die Serum-Halbwertszeit von gereinigter Protease in der Maus betrug etwa 60 nun. Ein Anstieg korrespondierender Antikörper war in alien infizierten Tieren zu beobachten. Auch im Serum gesunder Probanden waren Antiprotease-Antikörper bis zu einem Titer von 1:8000 nachweisbar; der Protease-Antigentest fiel hingegen immer negativ aus. Die Titer von Antikörpern gegen metabolische Candida-Antigene erreichten in derselben Gruppe von Seren Werte von 1:5120 bzw. 1:1280. Die sechs Tests wurden unter diagnostischen Bedingungen verglichen; Ergebnisse dieser Studie sind Gegenstand einer weiteren Mitteilung.     

血清唾液酸对血液病临床价值的初步报告

报道64例各类血液病患血清唾液酸（SA）值的测定，并与84例健康对照组进行比较。结果经统计学分析：病例组（631．5±130．3μg/ml），病例组中的ANLL（672．6±98．38μg/ml），ALL（630．8±29．3μg/ml），CML－A（617．5±122．7μg/ml），MM（606．0±138．7μg/ml四组与对照组比较，差异有极显意义（P＜0．01）；各病型组SA阳性率明显高于健康对照组，表明SA值的测定对恶性血液病的诊断有一定的临床意义。     

14岁以下正常人群B19病毒抗体的测定

用间接免疫荧光法检测了延边地区１４岁以下不同年龄组的正常人群血清中抗Ｂ１９病毒抗体。结果如下：抗体阳性率为１２．９％，各年龄组抗体阳性率分布不均，主要在７．５％～２２．５％之间，几何平均效价为１：１９，各年龄组的几何平均效价在１：１６～１：２５之间；抗体效价分布在１：１０～１：８０之间，主要在１：４０以下，朝鲜族和汉族的病毒抗体阳性率分别为９．７％和１６．０％，几何平均效价分别为１：２１和１：１８．统计结果表明，各年龄组之间及朝鲜族和汉族之间，抗体阳性率和几何平均效价均无显著性差异（Ｐ＞０．０５）．     

人巨细胞病毒pp150基因的原核表达及抗原性分析

①目的 构建人巨细胞病毒（HCMV）PP150的原核高效表达系统，制备用于急性，活动性HCMV感染血清学诊断的基因工程抗原。②方法 PCR扩增HCMV PP150抗原决定簇（840-1048aa）编码基因片段，分别插入原核表达载体PMAL-p2,pet-28a，转化宿主菌E.coli.诱导表达，经麦芽糖亲和层析树脂纯化，以WesternBlot及间接ELISA法鉴定及其抗原性。③结果 重组表达质粒PMAL-P2-PP150可在E.coli.5DH5a中有效表达，表达产物经SDS-PAGE电泳后，凝胶成像系统分析显示相对分子质量约为6.4万，约占菌体蛋白的10%。而重组质粒pET-28a-pp150未见明显表达带，Western-Blot检测，阳性识别率为80%（12/15）。用此蛋白包被ELISA板，检测正常孕妇血清，阳性率为6.5%(17/263)。与本室制备的全病毒抗原的ELISA的诊断符合率为96%。④结论此重组蛋白具有良好的抗原性，进一步完善后可用于HCMV急性和活动性感染的诊断。     

Serodiagnosis of tuberculosis and leprosy by enzyme immunoassay

Objective: To evaluate the use of serodiagnosis for tuberculosis and leprosy using mycobacterial antigen 38 kDa, with kits from Omega laboratories, to detect IgG by enzyme immunoassay (EIA).
Method: The study population consisted of 58 patients with evidence of tuberculous infection (culture of Mycobacterium tuberculosis complex or microscopic evidence), of whom 23 had pulmonary and 35 had extrapulmonary disease. There were six subjects who had recently been treated for tuberculosis, 11 patients on treatment for leprosy and 137 patients suspected of having tuberculosis on clinical or radiologic grounds (without laboratory evidence). A control group comprised 35 healthy individuals or patients suffering from diseases other than tuberculosis.
Results: The tests showed that there was a significant difference in antibody levels between the patients with active pulmonary disease, extrapulmonary tuberculosis and leprosy in comparison with the control group ( p <0.001). The sensitivities of the two tests together for proven pulmonary tuberculosis were 100% and 95.7% at 1.0–1.5 and >1.6 EIA cut-off points respectively, while the specificities were 88.5% and 100% at the same cut-off points. The sensitivities for extrapulmonary tuberculosis were 71.4% and only 51.4% at 1.0–1.5 and >1.6 EIA cut-off points. The test was positive in 30 (21.9%) of the 137 suspected patients, while 43 (31.4%) had an equivocal result and the remaining 64 (47.7%) suspects were definitely negative. There was again a significant difference in positivity rates between suspects and the control group.
Conclusions: Omega IgG test is useful in the serodiagnosis of active pulmonary tuberculosis and leprosy, but less sensitive in extrapulmonary disease, particularly in children. Equivocal results may only add to the evidence of tuberculosis in early or minimal disease.     

Kaposi's sarcoma-associated herpesvirus serology in Europe and Uganda: multicentre study with multiple and novel assays

A multicentre study was undertaken to define novel assays with increased inter-assay concordance, sensitivity, specificity and predictive value for serological diagnosis of human herpesvirus type 8 (HHV-8) infection. A total of 562 sera from European and Ugandan human immunodeficiency virus (HIV)-infected or uninfected individuals with or without Kaposi's sarcoma (KS) and blood donors were examined under code by 18 different assays in seven European laboratories. Sera from KS patients and all non-KS sera found positive by at least 70%, 80%, or 90% of the assays were considered "true positive." The validity of the assays was then evaluated by univariate logistic regression analysis. Two immunofluorescence assays (IFA) for detection of antibodies against HHV-8 lytic (Rlyt) or latent (LLANA) antigens and two enzyme-linked-immunosorbent assays (ELISA) (M2, EK8.1) for detection of antibodies against HHV-8 structural proteins were found to be highly concordant, specific, and sensitive, with odds ratios that indicated a high predictive value. When used together, the two IFA (Rlyt-LLANA) showed the best combination of sensitivity (89.1%) and specificity (94.9%). The performance of these assays indicate that they may be used for the clinical management of individuals at risk of developing HHV-8 associated tumours such as allograft recipients.     

Cell cultures for diagnosis of arbovirus infections in livestock and wildlife

Summary Arboviruses can be isolated in serially propagated cells derived from various vertebrates and invertebrates. Cell cultures can be used for direct detection of antigen by fluorescent antibody and enzyme-linked immunosorbent assays, for nucleic acid hybridization, and for visualization of viruses with electron microscopy. Reagents for enzyme-linked immunosorbent assays for IgM and IgG antibodies, hemagglutination-inhibition, complement fixation, and serum dilution-plaque reduction neutralization tests can be prepared in cell cultures infected with these viruses. Thus, cell cultures can be used as laboratory hosts for essentially all isolation, identification, and serodiagnostic procedures for arboviruses. This paper outlines current methods for diagnosis of arbovirus infections in livestock and wildlife, describes certain of these techniques, and provides references for others.     

体外免疫制备抗大肠癌循环抗原的单克隆抗体

用带结肠癌HRT-18细胞株的BALB/c(nu/nu)小鼠的血清,体处免疫BALB/c小鼠的脾细胞.再与Sp2/0细胞融合,经免疫荧光法在人结肠癌石蜡切片上筛选出一组抗结肠癌的单克隆抗体;A15-6,C13-11,H16-8。间接免疫酶法显示这组单抗对结肠癌的阳性率为69％-72％。免疫组化的特点为:癌巢分泌物及其接触的细胞膜顶端多为阳性反应,其他部位呈阴性反应。3抗体对其他类型的组织无反应。可见,这是一组针对血液循环中肿瘤相关抗原的单克隆抗体,有较好的器官特异性,可能有益于大肠癌的临床血清学检测。     

血管内皮生长因子在常见恶性肿瘤诊治中的临床价值

目的:探讨血管内皮生长因子(VEGF)在常见恶性肿瘤诊断、治疗和预后中的临床意义。方法:应用酶联免疫吸附法(ELISA)检测544例恶性肿瘤患者及87例对照者血清中VEGF浓度和CEA、CA50等6种肿瘤标记物的水平。计算VEGF的灵敏度、特异度、阳性预测值和阴性预测值,分析患者治疗前血清VEGF水平对其近期疗效的影响以及VEGF和其他肿瘤标记物联合检测结果与患者疗效的关系。结果:①各种肿瘤患者血清VEGF平均水平均高于对照组,其中以胃癌、肝癌和肺癌患者较高;②VEGF以200.6(ng/L)为医学参考值上限,其灵敏度为54.2%,特异度为95.4%;有负荷组患者灵敏度可达到74.9%,而无负荷者仅为20.4%,其差别有统计学意义(P<0.01);③肿瘤患者随着血清VEGF平均水平的升高,其疗效逐渐降低(P<0.05)。五种肿瘤标记物联合分析显示,随着患者血清肿瘤标记物阳性数目的增多,无效患者的比例呈上升趋势,当患者血清肿瘤标记物阳性数目达到3种或以上时,约89%的患者显示无效。结论:血清VEGF是一种广谱的肿瘤标记物,对多种肿瘤的辅助诊断均有重要的临床价值,且对初诊肿瘤患者,VEGF是一个良好的检测指标,其鉴别诊断价值优于其他6种肿瘤标记物。VEGF与其他肿瘤标记物联合检测有助于临床疗效评价、预后评估和病情随访监测。     

International multi-centre evaluation of a dipstick assay for human leptospirosis

A dipstick assay for the detection of Leptospira-specific immunoglobulin M (IgM) antibodies in human sera was evaluated in 27 laboratories in 23 countries. 873 serum samples from 711 patients including 329 laboratory-confirmed leptospirosis case patients, 239 noncase patients and 69 patients with viral infections causing heamorrhagic fever were tested. Relative to the results of the reference leptospirosis test, the sensitivity of the dipstick assay was 84.5% for serum samples collected during the first 10 days of the disease and 92.1% for serum samples collected 10-30 days after the onset of disease. The specificity was 87.5% and 94.4%, respectively. Similar to viral haemorrhagic fevers, leptospirosis may cause bleeding. A small number of serum samples from patients with haemorrhagic viral infections gave a weak (1 +) stain. All other samples were negative. In conclusion, the dipstick assay is sensitive and specific and reacts well with serum samples from patients infected with a range of leptospiral strains. It is also easy to use and does not require special equipment or refrigeration. Therefore the assay is ideal for use in developing countries and rural settings.