胶体金联免疫吸附试验在人免疫缺陷病毒抗体检测中…

将基因工程表达的人免疫缺陷病毒１型ｇｐ４１和２型ｇｐ３６作为抗原结合在硝基纤维素膜上，以四氯金酸结合的羊抗人ＩｇＧ作为标记抗体，建立了检测人血清中ＨＩＶ－１／２型抗体的胶体金联免疫吸附试验。ＧＣＩＳＡ检测抗体的特异性和敏感性均达到国内外酶联免疫吸附试验检测水平，并完全达到对ＨＩＶ－１／２型抗体诊断试剂国家质控标准的要求。     

人类免疫缺陷病毒基因变异的临床意义

人类免疫缺陷病毒（ＨＩＶ）基因变异是影响感染者抗病毒免疫反应的主要因素，同时也是影响抗ＨＩＶ疫苗发展的主要障碍，认识ＨＩＶ基因变异的规律将有助于抗ＨＩＶ药物治疗的临床应用。     

4种国产和3种进口第4代HIV抗原抗体检测试剂的质量评价

目的 对4种国产和3种进口第4代HIV诊断试剂的质量进行评价.方法 利用HIV抗体阴性样品库和核酸阳性样品库、BBI阳转血清盘样品等,对4种国产和3种进口第4代试剂的敏感性和特异性、检测HIV-1早期感染的能力进行分析.结果 7种第4代试剂的敏感性均为100％ (95％ CI:99.86％～100％),且1份HIV-1感染窗口期样品均检测为阳性,其中1种进口试剂“8+”值较大(1.0892),其余6种试剂的“δ+”值均比较小(0.0836 ～0.3003).对阴性样品,7种试剂均存在不同程度的假阳性(特异性为97.80％ ～ 99.60％,“δ-”值为-1.3803 -0.4778).对BBI阳转血清盘样品,国产试剂的阳转血清相对敏感性系数为-0.500～0,2种进口试剂则为-0.600和-0.700.结论 7种试剂均具有较高的敏感性和特异性,第4代HIV试剂用于血液筛查可发现HIV感染窗口期样本,对减少HIV传播的风险有一定的意义.但进口第4代试剂检测HIV-1早期感染的能力强于国产第4代试剂.     

人源抗人免疫缺陷病毒1型噬菌体抗体库的构建及筛选

目的构建人免疫缺陷病毒１型（ＨＩＶ－１）特异性噬菌体抗体库，制备人源抗ＨＩＶ－１ｇｐ１２０单克隆抗体。方法以半巢式聚合酶链反应（ＰＣＲ）从ＨＩＶ－１感染者外周血单个核淋巴细胞中扩增抗体重链Ｆｄ和轻链（ｋ）基因，与噬菌体载体ｐＣｏｍｂ３连接，构建噬菌体抗体Ｆａｂ组合文库。对抗体库进行３轮吸附－洗脱－扩增的亲和选择后，以ＥＬＩＳＡ法筛选抗ＨＩＶ－１ｇｐ１２０噬菌体抗体，并进行ＤＮＡ序列分析和Ｆａｂ的可溶性表达。结果半巢式ＰＣＲ有效地扩增出Ｆｄ和ｋ基因，以此构建成容量为１９５×１０７的噬菌体抗体库。３轮亲和选择使特异性抗体得到高度富集，抗ＨＩＶ－１ｇｐ１２０噬菌体抗体阳性克隆占３２％。对一阳性克隆抗体基因ＣＨ１和ＣＬ部分ＤＮＡ序列进行了测定，并在大肠杆菌表达出可溶性Ｆａｂ。结论抗ＨＩＶ－１特异性噬菌体抗体库的构建和人源抗ＨＩＶ－１ｇｐ１２０单克隆抗体的制备为今后筛选抗ＨＩＶ中和抗体奠定了基础，具有重要的应用价值。     

IL-16抗人免疫缺陷病毒的分子机制

IL 16主要来自CD8+ T淋巴细胞 ,它的受体是CD4分子。近来许多实验证实IL 16在体外有抑制人免疫缺陷病毒增殖的生物活性。其作用机制一为通过对抗NF κB和Tat所致的活化转录作用 ,从而影响人免疫缺陷病毒启动子的活性而抑制免疫缺陷病毒的转录。另一机制为下调CD95的表达 ,抑制T淋巴细胞活化后诱导的T淋巴细胞凋亡 ,从而增加T淋巴细胞的数目 ,为免疫缺陷病毒感染者的综合治疗提供了新的途径。     

1995年云南瑞丽人免疫缺陷病毒的生物学特性

１９９５年从云南省瑞丽市３０名静脉注射毒品者（ＩＤＵｓ）及部分配偶中采血，分离外周血单个核细胞（ＰＢＭＣ），用共培养法分离人免疫缺陷病毒（ＨＩＶ），以ＨＩＶ－Ｐ２４抗原酶联免疫吸附试验（ＥＬＩＳＡ）为检测终点，分离到１０株ＨＩＶ。其毒株的生物学特征为：复制能力弱，生长缓慢，滴度不高；不引起细胞融合。多数毒株只能在ＰＢＭＣ中生长，不能感染Ｔ细胞，只有１株（Ｃｒ２６９）可感染Ｔ细胞。已分离毒株的核苷酸     

抗人免疫缺陷病毒1型gp120人源单链抗体的表达

目的研究人源抗人免疫缺陷病毒１型（ＨＩＶ－１）ｇｐ１２０单链抗体（ＳｃＦｖ）。方法以人工合成的ＨＩＶ－ｌｇｐ１２０Ｖ３环多肽为抗原，利用噬菌体抗体库技术，筛选含有抗－ｇｐ１２０ＳｃＦｖ基因的噬菌体，提取质粒，转化大肠杆菌ＨＢ２１５１，表达可溶性ＳｃＦｖ。结果经ＳＤＳ－ＰＡＧＥ和Ｗｅｓｔｅｒｎｂｌｏｔ分析，表达产物分子量为２８ｋＤ左右，且具有ｃ－ｍｙｃ活性；ＥＬＩＳＡ和Ｄｏｔｂｌｏｔ结果表明，可溶性ＳｃＦｖ具有较好的抗原结合活性和较强的特异性；竞争性ＥＬＩＳＡ实验结果进一步证明表达产物的特异抗－ｇｐ１２０活性。结论该技术便捷有效，可大量获得人源抗ＨＩＶ抗体片段，为进一步研究抗ＨＩＶ抗体的生物活性和ＨＩＶ感染诊治打下基础     

HIV-1/2抗体和P24抗原联合检测试剂盒的研制与评价

目的研制HIV-1／2抗体和P24抗原联合检测酶免疫试剂盒并评价其实用性。方法联合使用基因工程HIVI／2型抗原和抗HIVP24单克隆抗体包被酶联反应板，以辣根过氧化物酶标记的HIVI／2型抗原和生物素化的兔抗HIVP24抗体作为标记物，研制了联合检测HIVI／2抗体和P24抗原的ELISA诊断试剂，并对其特异性、敏感性、稳定性等进行评价和临床考评。结果检测P24抗原质控品的灵敏度可达0．2ng／ml；与雅培公司试剂比较检测78份AIDS患者血清和85份正常人血清、对照检测中国药品生物制品检定所研制的HIV参比血清，特异度和灵敏度均为100％。临床考核检测12051份各种血清，灵敏度为100％（543／543），特异度为99．48％（11448／11508）。试剂在37℃放置6d后，试验结果无明显差异。结论本试剂盒具备特异度强、敏感度高、稳定性好、操作简便等优点，可以一步检出HIV特异性抗体和HIVP24抗原，缩短了HIV感染的检测窗口期，适用于HIV感染的实验室诊断和流行病学调查。     

HIV抗体酶联免疫诊断试剂检测不同基因型抗体的研究

目的 分析不同HIV抗体酶联免疫诊断试剂对检测HIV不同基因型抗体的情况。方法 对不同地区的20份HIV抗体阳性样品中HIV核酸进行扩增，对PCR产物进行测序并进行基因型别分析。用不同试剂对系列稀释的不同基因型样品进行检测。结果 20份样品均为HIV RNA阳性，其中9份样品为HIV B亚型，9份样品为：HIV C或BC重组，2份为HIVAE重组。不同试剂对HIV不同基因型抗体的检测灵敏度无明显差异。结论 我国主要的商业化HIV抗体诊断试剂产品检测不同基因型抗体的能力无明显差异。     

Relevance of cutoff on a 4th generation ELISA performance in the false positive rate during HIV diagnostic in a low HIV prevalence setting

BackgroundDespite the high specificity of fourth-generation enzyme immunoassays (4th-gen-EIA) for screening during HIV diagnosis, their positive predictive value is low in populations with low HIV prevalence. Thus, screening should be optimized to reduce false positive results.ObjectivesThe influence of sample cutoff (S/CO) values by a 4th-gen-EIA with the false positive rate during the routine HIV diagnosis in a low HIV prevalence population was evaluated.Study designA total of 30,201 sera were tested for HIV diagnosis using Abbott Architect^® HIV-Ag/Ab-Combo 4th-gen-EIA at a hospital in Spain during 17 months. Architect S/CO values were recorded, comparing the HIV-1 positive results following Architect interpretation (S/CO ≥ 1) with the final HIV-1 diagnosis by confirmatory tests (line immunoassay, LIA and/or nucleic acid test, NAT). ROC curve was also performed.ResultsAmong the 30,201 HIV performed tests, 256 (0.85%) were positive according to Architect interpretation (S/CO ≥ 1) but only 229 (0.76%) were definitively HIV-1 positive after LIA and/or NAT. Thus, 27 (10.5%) of 256 samples with S/CO ≥ 1 by Architect were false positive diagnose. The false positive rate decreased when the S/CO ratio increased. All 19 samples with S/CO ≤10 were false positives and all 220 with S/CO > 50 true HIV-positives. The optimal S/CO cutoff value provided by ROC curves was 32.7. No false negative results were found.ConclusionsWe show that very low S/CO values during HIV-1 screening using Architect can result HIV negative after confirmation by LIA and NAT. The false positive rate is reduced when S/CO increases.     

Comparative evaluation of three FDA-approved HIV Ag/Ab combination tests using a genetically diverse HIV panel and diagnostic specimens

BackgroundHIV Ag/Ab combination assays are recommended by CDC for routine screening and several HIV Ag/Ab combination tests are now FDA-approved. Maintaining high specificity and consistent sensitivity across diverse HIV strains is critical for these assays to accurately detect HIV infection and expedite delivery of patient results.ObjectivesTo evaluate performance of three FDA-approved HIV tests: ARCHITECT HIV Combo (Abbott), ADVIA Centaur HIV Combo (Siemens) and BioPlex HIV Ag-Ab (Bio-Rad).Study designSensitivity and specificity were evaluated using an extensive panel of 28 HIV infected human specimens and 17 cultured virus isolates representing multiple genotypes, 6 seroconversion panels, 4 human samples with acute infection, WHO p24 standard and 4020 clinical specimens.ResultsThe p24 limit of detection (LOD) for the WHO standard was 0.19 IU/ml, 0.70 IU/ml, and 1.77 IU/ml in BioPlex, ARCHITECT, and Centaur respectively. The distribution of LODs across 15 HIV-1 isolates was substantially narrower in ARCHITECT (5–33 pg/ml) than in BioPlex (11–198 pg/ml) and Centaur (6–384 pg/ml). All assays detected antibodies to the majority of HIV-1 and HIV-2 variants. However, reduced sensitivity was observed for Centaur in detection of antibodies to HIV-1 group M (CRF02_AG), O and N variants. BioPlex and ARCHITECT showed better seroconversion sensitivity than Centaur, detecting one bleed (3–7 days) earlier in 4 (BioPlex) and 3 (ARCHITECT) of 6 seroconversion panels. ARCHITECT demonstrated the highest specificity (99.90–100%) compared to BioPlex (99.80%) and Centaur (99.42%).ConclusionsThe overall performance of ARCHITECT and BioPlex was superior to Centaur, especially for detection of acute HIV infection.     

Detectability index,likelihood ratio and diagnostic output as psychophysical measures for evaluation of medical image processing

Signal-detection theory was used to study methods of measuring changes in diagnostic quality from medical image processing. Detectability index and likelihood ratio are applicable for purposes of comparison but they do not give an indication of the confidence with which a diagnosis has been made. Therefore a new measure for evaluation, the diagnostic output, is defined which incorporates this important aspect. This study describes experiments where four observers judge pictures of heart scintigrams in different stages of processing involving variations of resolution and brightness. The emphasis is on methods for evaluation of the diagnostic value of picture processing. Our conclusion is that the influence of the studied image-processing techniques can be well described using signaldetection theory, while the diagnostic output seems to be a well suited measure to represent diagnostic properties of stimulus and observer.     

四种乙肝病毒表面抗原ELISA诊断试剂的评价

目的 对四种国内乙肝病毒表面抗原诊断试剂进行评价。方法 用四种试剂分别对标准品和未知血清进行检测，用几种常用诊断试验评价方法对检测结果进行分析和比较。结果 四种试剂对标准血清的检测结果较好，A、B两种试剂的总符合率为100％，另两种的总符合率也在90％以上，其中B试剂有较好的精密性。δ值比较显示A和B两种试剂有较好的诊断特性。四种试剂在一定的抗原浓度范围内都有良好的线性关系。针对于未知样本的检测中，四种试剂与采用电发光法的罗氏诊断试剂有较好的一致性。结论 国内主要厂家的HBsAg ELISA诊断试剂具有较高的检测效能。在实际工作中，可以通过常用的诊断试剂筛选实验，经统计分析后筛选出较高效能诊断试剂用于检测。     

Contrast agents for diagnostic ultrasound: development and evaluation of polymer-coated microbubbles

Although the concept of an ultrasound contrast agent dates from Gramiak's work in 1968 in which indocyanine green was injected into the ascending aorta and heart, no universally accepted contrast agent for ultrasound now exists. This is primarily due to problems with stability, size and/or toxicity of the agents which have been investigated. Development of an effective ultrasound contrast agent would be highly significant for the health care industry, since it would greatly expand the scope of ultrasound (a noninvasive and safe procedure) as a diagnostic technique. While encapsulated gas bubbles offer particular advantages in stability over hand-agitated systems, they frequently present problems with size. Capsules larger than 10 μm in diameter become entrapped in the capillary bed of the lung. This paper describes the use of ionotropic gelation of the naturally occurring polysaccharide, alginate, for microencapsulation of air. Two procedures have been investigated. A novel jet head has been developed which allows co-extrusion of a solution of sodium alginate and air to produce nascent microencapsulated air bubbles which fall into a hardening solution of calcium ions. A second method employs ultrasound to introduce cavitation-induced bubbles into the alginate before capsule formation by spraying. Power spectra of these preparations demonstrate echogenicity (that is strong scatter of the incident ultrasound wave back to the emitting transducer, which also acts as a receiver), with resonant peaks that are a function of capsule size and wall characteristics.     

Economic evaluation of point-of-care diagnostic technologies for infectious diseases

We review the growing number of economic evaluations of individual point-of-care (POC) tests for diagnosis of infectious diseases in resource-limited settings that use either cohort studies or mathematical models. We focus on studies that evaluate POC diagnostic tests for the control of human immunodeficiency virus (HIV) and malaria, tools that are central to the WHO prevention guidelines for infectious diseases in developing countries. Although rapid diagnostic tests for HIV and malaria seem to be cost-effective in these standard analyses, these do not take into account the reduction in patients’ waiting time and the number of clinic visits required to receive results, or future benefits from the reduction in antimalarial drug pressure. Those additional cost reductions would be considerably greater with POC rapid tests, and the cost-effectiveness of POC tests would therefore be improved. Findings from cost-effectiveness analyses suggest that, despite the relatively small additional cost incurred, decision-makers should strongly consider using POC tests throughout or during parts of HIV and malaria epidemics, where this is feasible in terms of local human resources and logistical conditions.     

The fourth generation AlereTM HIV Combo rapid test improves detection of acute infection in MTN-003 (VOICE) samples

BackgroundEarly and accurate detection of HIV is crucial when using pre-exposure prophylaxis (PrEP) for HIV prevention to avoid PrEP initiation in acutely infected individuals and to minimize the risk of drug resistance in individuals with breakthrough infection.ObjectiveTo determine if fourth-generation antigen/antibody (Ag/Ab) rapid diagnostic tests (RDT) would have detected HIV infection earlier than the third-generation RDT used in MTN-003 (VOICE).Study design5029 VOICE participants were evaluated with third-generation Alere Determine™ HIV-1/2, OraQuick ADVANCE^® Rapid HIV-1/2, Uni-Gold™ Recombigen^® HIV-1/2 and Bio-Rad GS HIV-1/2 + O EIA; and fourth-generation Alere Determine™ HIV-1/2 Ag/Ab Combo, Conformité Européene (CE)-Marked Alere™ HIV Combo and Bio-Rad HIV Combo Ag/Ab EIA. Multispot^®, GS HIV-1 Western Blot (WB) and Geenius™ (Bio-Rad) were also evaluated.ResultsOf 57 antibody-negative pre-seroconversion plasma samples with HIV RNA >20 copies/mL identified, 16 (28%) were reactive by CE-Marked Alere™ HIV Combo (1 Ab; 9 Ag; 6 Ag/Ab reactive) and 4 (7%) by Alere Determine™ HIV-1/2 Ag/Ab Combo (2 Ab; 2 Ag; 0 Ag/Ab reactive) (p = 0.0005). Multispot^® confirmed only 1 of 16 acute infections while WB and Geenius™ confirmed none. GS HIV Combo Ag/Ab EIA identified 27 of 57 (47%) pre-seroconversion RNA-positive samples.ConclusionIn VOICE, 28% of infections missed by current third-generation RDT would have been identified with the use of CE-Marked Alere™ HIV Combo. Geenius™, Multispot^® and WB were all insensitive ( < 10%) in confirming infections detected by fourth-generation assays. An improved diagnostic algorithm that includes a fourth-generation RDT with HIV RNA testing will be essential for efficiently identifying seroconverters on PrEP.