Evaluation of a new Trypanosoma cruzi antibody assay for blood donor screening

BACKGROUND: This multicenter prospective study was designed to evaluate the performance characteristics of a new commercially available enzyme-linked immunosorbent assay (ELISA) for the detection of antibodies to Trypanosoma cruzi in blood donors, the ORTHO T. cruzi ELISA Test System (Ortho-Clinical Diagnostics). STUDY DESIGN AND METHODS: Assay specificity was evaluated among 40,665 serum and ethylenediaminetetraacetate (EDTA) plasma specimens from volunteer blood donors and 481 T. cruzi antibody-negative specimens from a high-risk population. Sensitivity was evaluated among 106 T. cruzi-infected subjects identified by parasite detection, among 93 radioimmunoprecipitation assay (RIPA)-positive specimens from high-risk subjects, and 662 specimens presumed positive for the presence of T. cruzi antibodies by serologic methods. Also assessed were the equivalence of serum and plasma as specimen sources, performance equivalence of automated and semiautomated processing methods, nonspecific reactivity in specimens from other disease states or clinical conditions, and assay precision. RESULTS: Assay specificity was 99.998 percent in volunteer blood donors and 99.4 percent among high-risk subjects. Sensitivity was 100 percent among specimens positive by parasite detection, or by serologic methods, and 98.9 percent among RIPA-positive specimens from high-risk subjects. No differences were demonstrated between serum and plasma or between semiautomated and automated processing methods. Cross-reactivity was observed with known positive leishmaniasis specimens. Total inter- and intraassay variability was less than 10 percent with both the automated and the semiautomated methods. CONCLUSION: The ORTHO T. cruzi ELISA Test System is an effective, qualitative assay for screening blood donors for immunoglobulin G antibodies to T. cruzi. The assay was licensed for donor screening by the FDA in December 2006.     

Combination HCV core antigen and antibody assay on a fully automated chemiluminescence analyzer

BACKGROUND: HCV exposure among blood donors is serologically determined by detection of antibodies to HCV (anti-HCV); however, the recent development of an assay for the detection of HCV core antigen identifies infection before anti-HCV development. Simultaneous detection of HCV core antigen and anti-HCV would shorten the window period before seroconversion over conventional HCV antibody screening assays. STUDY DESIGN AND METHODS: A prototype chemiluminescent immunoassay was developed for simultaneous detection of HCV core antigen and anti-HCV in human sera and plasma. The assay was performed on a single-channel instrument representing an automated serologic analyzer (PRISM, Abbott Laboratories) system. Sensitivity and specificity were evaluated by testing 23 HCV seroconversion panels and plasma or sera from volunteer blood donors. RESULTS: The prototype HCV core antigen and antibody combination assay detected 80 of 89 (89.9% ) HCV RNA-positive and antibody-negative specimens from 23 panels, thereby reducing the seroconversion window period by an average of 34.3 days compared to PRISM HCV antibody detection. All PRISM HCV antibody-positive specimens were detected by the combination assay for a relative sensitivity of 100 percent. The repeatedly reactive rate was 0.20 percent based on testing of 3017 screened anti-HCV-negative sera and plasma. CONCLUSIONS: The prototype combination assay was shown to detect HCV core antigen and anti-HCV simultaneously and significantly closed the time gap between the initial detection of HCV RNA and the first appearance of detectable antibodies to HCV.     

全自动血型分析仪应用于献血者血型筛查

目的探讨并评价全自动血型分析仪应用于献血者血型筛查和盐水不规则抗体检测。方法采用全自动血型分析仪(全自动法)对25 554例献血者标本作ABO及RhD血型鉴定、盐水不规则抗体初筛,并与加样仪加样手工比色法(半自动法)作比对实验。ABO正反定型不一致而无法定型、O细胞凝集、RhD阴性的标本送血型红细胞参比实验室鉴定。结果全自动法与半自动法比较,ABO、RhD阴性血型1次准确定型率:99.93%(25 535/25 554)vs99.95%(25 542/25 554)(P>0.05);O细胞凝集阳性率:0.18%(46/25 554)vs0.10%(26/25 554)(P<0.05),经参比实验室确认,盐水不规则抗体分别为43、24例(0.17%vs0.09%,P<0.05);正反定型不符:17例(0.06%)vs10例(0.04%),参比实验室确认,经参比实验室确认2种方法共同拥有亚型5例,亚型漏检各2例(0.01%),余为正常血型(10/17vs3/10,P>0.05)。结论全自动血型分析仪作ABO、RhD血型筛查的技术相关性好、重复性好;除了全自动化、操作规范化、标准化等优点外,全自动血型分析仪更易发现盐水不规则抗体。     

Evaluation of a fully automated procalcitonin chemiluminescence immunoassay

We evaluated a new fully automated microparticle immunoassay for procalcitonin (LIAISON BRAHMS PCT) in comparison with a previously established manual chemiluminescence assay from the same manufacturer (LUMItest PCT, BRAHMS AG). Procalcitonin (PCT) is an early and rather specific marker of systemic bacterial infection. In addition, the efficacy of antibiotic therapy can be monitored by sequential analysis of PCT values. This is why rapid and accurate determinations of PCT are urgently required by intensive care units. The aim of this study was to evaluate in a clinical set-up a new fully automated rapid PCT test. Analytical results are compared with results obtained by a previously introduced quantitative manual test. Intra-assay coefficients of variation (CV) were found in the range of 0.94 to 7.1% at concentrations between 0.46 and 97.2 microg/l. Over a time period of 27 days the inter-assay CV was found below 4.0% at concentrations of 1.93 and 14.29 microg/l and 9.9% at 0.40 microg/l. The functional sensitivity at a CV level of 20% was determined as 0.2 microg/l. Linearity could be demonstrated in a concentration range from 0 to 445 microg/l. When serum and plasma with EDTA, citrate or heparin anti-coagulation were analyzed in parallel, no systematic bias was found. A method comparison by regression analysis showed PCT values determined by both tests in very good agreement (r = 0.99). PCT concentrations in apparently healthy subjects (n =101) were below 0.58 microg/l in line with previously published results. Patients with sepsis (n = 43) or with infectious adult respiratory distress syndrome (ARDS) (n = 28) showed median values of 22.2 and 18.9 microg/l, respectively. In a clinical set-up the LIAISON Brahms PCT assay provided rapid and accurate PCT results supporting the early detection of severe sepsis, the differentiation between systemic bacterial infection and other inflammatory diseases, and the monitoring of antibiotic therapy in septic patients. The results of the new LIAISON BRAHMS PCT assay show an excellent concordance with the LUMItest PCT. The clinical information derived from the measurements is well comparable to the results obtained with the LUMItest PCT, too.     

Evaluation of a new fully automated one-step C-peptide chemiluminescence assay (LIAISON C-Peptid)

The determination of C-peptide, a 31 amino acid fragment of proinsulin which is a byproduct of insulin formation, is used as a marker for insulin secretion. Clinically, the determinations are performed to detect autonomous insulinoma, factitious hypoglycemia, and in general to assess the function of beta-cells in patients with diabetes mellitus. The analysis is frequently performed by radioimmunoassays (RIA), which have several disadvantages, for instance the use of radioactivity and time and resource requirements. We performed an evaluation of a new fully automated chemiluminescence assay (LIAISON C-Peptid, Byk-Sangtec) at two clinical sites, in Germany and Italy, with regard to imprecision and clinical relevance of the obtained data, and the correlation with a standard RIA method and another chemiluminescence test. The new assay showed a good correlation with the RIA (r = 0.950) and the chemiluminescence assay (r = 0.967). The intra-assay variability and inter-assay variability was 3.5% and 8.7% in Germany, and 2.4% and 9.6% in Italy. The clinical evaluation of samples derived from 19 oral glucose tolerance tests, 13 insulin suppression tests, and 2 insulin secretion stimulation tests revealed a clinical specificity of 100%, i.e. all cases resulted in the same clinical diagnosis with all tests. With regard to the practical performance of the assays, the new chemiluminescence test, as a single-step fully automated method, offered the advantage of being a non-radioactive, less complex and much faster method than the RIA and also had timely advantages over the comparative chemiluminescence test. In general, the new LIAISON chemiluminescence assay compared favorably with the RIA and comparative chemiluminescence test and offers an attractive alternative for C-peptide analysis.     

迈克IS1200全自动化学发光测定仪的性能评价

目的对迈克IS1200全自动化学发光测定仪的性能进行评价。方法按照美国临床和实验室标准协会文件的要求,通过一系列实验设计,在IS1200上对乙肝5项、艾滋、丙肝和梅毒共8项进行检测,评价其精密度、试剂开瓶稳定性、线性和参考区间,与雅培i2000全自动化学发光分析仪的检测结果进行方法学比对。结果除乙型肝炎E抗原低值的总精密度略高于判断标准15%外,其余指标均符合要求;各项目的试剂开瓶稳定性良好;定量项目乙型肝炎表面抗体检测范围内的线性良好(r20.95);参考区间验证中,各项目均未发现离群值;方法学比对中,除丙型肝炎病毒抗体外,其他项目与雅培i2000全自动化学发光分析仪的相关性良好。结论迈克IS1200全自动化学发光测定仪的检测性能良好,满足实验室免疫检测工作的需求。     

Evaluation of a fully automated centrifugal analyzer for performance of hemostasis tests

We have evaluated the performance of a centrifugal analyzer, specifically designed for hemostasis tests, for methods based on clotting time as measured by light scattering, or based on splitting of chromogenic substrates. The results were compared with those obtained with a semiautomated coagulometer (clotting tests) or with a manually operated photometer (chromogenic tests). In general, the results with the centrifugal analyzer were at least as precise as those of the comparative methods, with greater output and ease of operation. Disadvantages of the instrument are its partial incompatibility with some commercial reagents and the relative rigidity of the operational parameters, which hinder its use for research.     

Evaluation of a new PCR assay with competitive internal control sequence for blood donor screening

BACKGROUND: High-throughput nucleic acid testing for transfusion-relevant viruses by PCR requires contamination-proof methods with high sensitivity and validity. A new PCR reagent kit (TaqMan, PE BioSystems) reduces the risk of carry-over contamination by eliminating post-PCR processing. STUDY DESIGN AND METHODS: Oligonucleotide design was done with software specialized for designing the assays' (TaqMan) primers and probes. A template-derived competitive internal control sequence designed through site-directed mutagenesis was used to reveal failures in amplification. Assay sensitivity was determined for single-donor and single-patient testing and by spiking sample mini-pools. Three seroconversion panels were tested. RESULTS: Sensitivity is high, reaching 300 HBV genomes per mL of single-patient material on direct testing. A detection limit of 1000 HBV genome equivalents per mL of donor plasma is achieved for 96 pooled samples. The window period for HBV infection was reduced by 17, 10, and 63 days from that for HBsAg screening in three seroconverting donors. CONCLUSION: The assay provides sufficient sensitivity to be superior to HBsAg screening in transfusion medicine and will be useful in clinical laboratories because of its ease of handling.     

An improved serodiagnostic test for Chagas' disease employing a mixture of Trypanosoma cruzi recombinant antigens

BACKGROUND: Blood transfusion is one of the most important transmission routes of Chagas' disease, a major parasitic infection in Latin America. Therefore, screening for antibodies to Trypanosoma cruzi is mandatory in blood banks in South America. Most of the commercial serologic tests employ epimastigote antigens and show a high number of inconclusive and false-positive results, with high economic and social costs. STUDY DESIGN AND METHODS: An ELISA using a mixture of three T. cruzi recombinant antigens, B13, 1F8, and H49 (mix-ELISA), was evaluated, first with a panel of well-characterized sera from 617 patients with Chagas' disease and 277 nonchagasic individuals, living in nine countries of South and Central America. Subsequently, the mix-ELISA was evaluated with 451 samples, from an endemic area of Brazil (Goiás), that were rejected from several blood banks because they presented discrepant results by two commercially available kits (indirect immunofluorescence assay, indirect hemagglutination assay, and/or ELISA). RESULTS: The mix-ELISA exhibited 99.7 percent sensitivity and 98.6 percent specificity in the first evaluation with the 894 samples. In the second evaluation, 451 sera that had discrepant results in the first screening for Chagas' disease were further analyzed with the mix-ELISA. Upon consideration of the consensus results obtained with the trypomastigote excreted-secreted antigens blot test, a confirmatory test for Chagas' disease, the mix-ELISA led to a reduction in 99.6 percent in the number of discordant sera. CONCLUSION: The combination of three T. cruzi recombinant antigens in a multiantigen immunoassay was highly sensitive and specific for Chagas' disease diagnosis. It is proposed that it can be applicable in blood bank screening in conjunction with the conventional serologic tests.     

Diana-5全自动血液分析仪白细胞分类与镜检的相关性研究

目的 对Diana-5全自动血液分析仪白细胞五分类与镜检的相关性进行研究与评价。方法 系统研究Diana-5全自动血液分析仪白细胞分类测定的精密度、总变异、镜检率、敏感性、特异性和过筛符合率，并分析比较与镜检结果的相关性。结果仪器分类测定中性粒细胞（Neu）、淋巴细胞（Lym）、单核细胞（Mon）、嗜酸性粒细胞（Eos）和嗜碱性粒细胞（Bas）的精密度分别为1.25％、1.53％、5．96％、7．58％和15．85％，总变异分别为1．82％、2．86％、7．84％、9．32％和17．32％。与镜检结果的相关系数分别为0．982、0．956、0．842、0．536和0．328。分类镜检率为20．2％，敏感性98．0％，特异性87．2％，过筛符合率88．1％。该仪器可提示异常白细胞的检测信息，其中非成熟细胞占异常提示信息的51.5％。结论 Diana-5血液分析仪白细胞分类测定的自动化程度高，结果 重复性好，具有良好的相关性，能有效发挥仪器的过筛作用，非常适合血常规的快速检测。     

Diana-5全自动血液分析仪白细胞分类与镜检的相关性研究

目的对Diana-5全自动血液分析仪白细胞五分类与镜检的相关性进行研究与评价。方法系统研究Diana-5全自动血液分析仪白细胞分类测定的精密度、总变异、镜检率、敏感性、特异性和过筛符合率,并分析比较与镜检结果的相关性。结果仪器分类测定中性粒细胞(Neu)、淋巴细胞(Lym)、单核细胞(Mon)、嗜酸性粒细胞(Eos)和嗜碱性粒细胞(Bas)的精密度分别为1.25%、1.53%、5.96%、7.58%和15.85%,总变异分别为1.82%、2.86%、7.84%、9.32%和17.32%。与镜检结果的相关系数分别为0.982、0.956、0.842、0.536和0.328。分类镜检率为20.2%,敏感性98.0%,特异性87.2%,过筛符合率88.1%。该仪器可提示异常白细胞的检测信息,其中非成熟细胞占异常提示信息的51.5%。结论Diana-5血液分析仪白细胞分类测定的自动化程度高,结果重复性好,具有良好的相关性,能有效发挥仪器的过筛作用,非常适合血常规的快速检测。     

Measuring calprotectin in plasma and blood with a fully automated turbidimetric assay

Calprotectin in plasma and blood might prove to be a useful biomarker of inflammation and infection; however, automated methods for analysing the concentration of calprotectin in those materials are lacking. We have validated a fully automated turbidimetric method and present health-related reference limits. Calprotectin was measured by Siemens Advia XPT with the Bühlmann fCAL® turbo test (Bühlmann Laboratories AG, Schönenbuch, Switzerland), a particle enhanced turbidimetric immunoassay for quantification of calprotectin in fecal extracts. Plasma and serum samples were analysed directly, while whole blood was first extracted with M-PER® Mammalian Protein Extraction Reagent (ThermoFisher) and diluted with B-CAL-EX (Bühlmann). We studied analytical imprecision, estimated health-related reference limits and examined the correlation between neutrophil-calprotectin (blood-calprotectin adjusted for plasma-calprotectin) and the neutrophil count. The intermediate (‘day-to-day’) coefficient of variation was 3.5 and 1.0% for heparin-plasma-calprotectin at 0.52?mg/L and 3.53?mg/L, respectively, and 4.9% for heparin-blood-calprotectin at 50.2?mg/L. Health-related reference limits were 0.470–3.02?mg/L for calprotectin in heparin-plasma, 50.8–182?mg/L for calprotectin in heparin-blood, 0.534–2.41% for the ratio between them and 24.7–33.3?pg for the mean amount of calprotectin per neutrophil. Compared to heparin-plasma, calprotectin concentrations were significantly lower in EDTA-plasma and higher in serum (p?<?.05). Correlation between neutrophil-calprotectin and the neutrophil count was excellent. We have shown that the Bühlmann fCAL® turbo test can be used to measure calprotectin in plasma and blood.     

Performance evaluation of the new fully automated human immunodeficiency virus antigen-antibody combination assay designed for blood screening

BACKGROUND: Before the introduction of human immunodeficiency virus (HIV) combination assays, serologic diagnosis of HIV infection was performed with assays that detected either antibodies or p24 antigen. Owing to the capability to detect the early appearance of p24 antigen, combination assays that are designed for simultaneous detection of antibodies and antigen can significantly reduce the diagnostic window. STUDY DESIGN AND METHODS: Specificity and sensitivity of a commercially available HIV antigen‐antibody combination assay (Abbott PRISM; assay is not licensed by the FDA for use in the United States) were evaluated in a multicenter study by testing volunteer blood donors, hospitalized patients, seroconversion panels, and p24 antigen and HIV antibody subtype panels. Performance data were compared to a commercially available HIV combination assay and the PRISM HIV O Plus assay. RESULTS: Apparent specificity of 99.95 percent was observed in the donor population for the PRISM HIV antigen‐antibody combination assay, and better seroconversion sensitivity was demonstrated compared with another combination assay and the PRISM HIV O Plus assay. Analytical HIV antigen detection sensitivity averaged 33 pg per mL on the Agence Française de Sécurité Sanitaire des Produits de Santé (AFSSAPS) panel. Furthermore, comparable antigen sensitivity was demonstrated for 32 HIV‐1 group M subtype and group O panels. The PRISM HIV combination assay detected all HIV‐1 group M and O and HIV‐2 antibody–positive specimens evaluated. CONCLUSIONS: The PRISM HIV antigen‐antibody combination assay demonstrated a significant reduction of the window period for diagnosis of HIV infection. The assay demonstrated enhanced specificity and sensitivity along with broad subtype detection. The assay performance represents the “state‐of‐the art” technology for serologic blood screening of HIV infection.     

化学发光法检测HBsAg在血液初筛中的应用评估

目的与酶联免疫吸附试验(ELISA)检测相比较,探讨化学发光法(CLIA)在血液初筛中的应用。方法采用1种CLIA和2种ELISA的乙肝表面抗原(HBs Ag)试剂平行检测日常献血者血样3 000份及卫生部临检中心收集的血清盘标本792份。任1种试剂有反应性的献血者标本和所有的血清盘标本由卫生部临检中心进行确认实验,分析比较结果。结果 3 000份献血者标本中11例有反应性,其中7份CLIA、ELISA和确认实验均为阳性; 4份为ELISA双试剂阴性CLIA单阳性,而确认结果均为阴性。792份血清盘标本中587份确认阳性,197份确认阴性,8份不确定。CLIA的HBs Ag最低检出浓度0. 05 IU/m L低于ELISA法的0. 09 IU/m L。CLIA对变异性抗原的检出率84. 62%明显高于ELISA B试剂42. 31%(P <0. 05)。CLIA的灵敏度89. 10%、特异性97. 97%明显高于2种ELISA法。重复性试验中,CLIA法的批内批间变异系数(CV)明显小于ELISA方法。结论 3 000份献血者血样经HBs Ag胶体金快速检测合格后抗原阳性率低,而792份血清盘由收集于不同血站的献血者标本组成,阳性率高,且标本的分组与设计合理有针对性。结合其检测结果,CLIA相比ELISA对HBs Ag的初筛有着较高的灵敏度、特异性和较好的精密度,且变异性抗原检出率较高,对防止不合格血样漏检,保证血液质量安全有较好的应用价值。     

UF-100尿液分析仪筛检尿路感染临床意义的探讨

目的　探讨UF 10 0尿液分析仪筛检尿路感染的临床意义。方法　对UF 10 0作重复性试验 ,用UF 10 0检测 32 5份尿中细菌和白细胞 ,同时作定量细菌培养并将结果作比较。用Yerushalmy模式评价 2种方法的一致性及UF 10 0筛检的灵敏度、特异性等。结果　重复试验中 ,UF 10 0细菌计数CV值低于白细胞计数CV ,与定量细菌培养结果比较 ,细菌计数的筛检灵敏度为 80 .0 % ,特异性为 5 0 .4 % ,阳性预计值为 2 8.7% ,阴性预计值为 91.0 % ,假阳性率为 39.7% ,假阴性率为 4 .0 % ,准确率为 5 6 .3%。结论　UF 10 0具有良好的分析尿液的性能 ,在临床尿路感染筛检时可用细菌一项指标 ,90 %结果阴性的标本可在短时间内筛去 ,大大减少实验人员繁复劳动 ,降低检验成本 ,但应注意假阴性 ,更不可替代尿定量细菌培养。     

UF-100尿液分析仪筛检尿路感染临床意义的探讨

目的 探讨UF-100尿液分析仪筛检尿路感染的临床意义。方法 对UF-100作重复性试验，用UF-100检测325份尿中细菌和白细胞，同时作定量细菌培养并将结果作比较。用Yerushalmy模式评价2种方法的一致性及UF-100筛检的灵敏度、特异性等。结果 重复试验中，UF-100细菌计数CV值低于白细胞计数CV，与定量细菌培养结果比较，细菌计数的筛检灵敏度为80．0％，特异性为50．4％，阳性预计值为28．7％，阴性预计值为91．0％，假阳性率为39．7％，假阴性率为4．0％，准确率为56．3％。结论 UF-100具有良好的分析尿液的性能，在临床尿路感染筛检时可用细菌一项指标，90％结果阴性的标本可在短时间内筛去，大大减少实验人员繁复劳动，降低检验成本，但应注意假阴性，更不可替代尿定量细菌培养。     

Evaluation of a low-ionic-strength solution-monospecific anti-IgG antiglobulin technique for donor antibody screening

Three different techniques of antibody screening of donor bloods were sequentially evaluated. Group I (16,300 donors) was a standard saline- albumin-AHG technique utilizing polyvalent serum, including incubation at room temperature. In Group II (26,243 donors), incubations (including room temperature) were performed in LISS, and monovalent anti-IgG serum was used. For Group III (15,840 donors), the room temperature incubation was not used for the LISS-IgG method of Group II. The three methods were comparable in terms of detection of clinically significant antibodies, while in Group III the detection of clinically nonsignificant antibodies was eliminated. Cost analysis indicates that for a donor center processing approximately 50,000 units per year and willing to prepare its own LISS solutions, conversion to LISS-IgG could produce a savings of between $5,000 and $8,000 per year. LISS-IgG is thus a sensitive and economical technique highly recommended for the donor center committed to manual donor antibody screening.     

Cost‐effectiveness of screening the US blood supply for Trypanosoma cruzi

BACKGROUND: Trypanosoma cruzi, the etiologic agent of Chagas disease, is a potential threat to transfusion recipients in the United States. The cost‐effectiveness of seven testing strategies was evaluated against no testing and hierarchically in incremental analysis. Donor‐specific strategies included testing donors born in endemic countries, testing all donors a specific number of times, or testing all donors every time. Component‐specific strategies are based on screening platelet‐containing donations. STUDY DESIGN AND METHODS: A decision analytic model simulated the lifetime cost (US dollars) and health outcomes (quality‐adjusted life‐years [QALYs]) of two hypothetical cohorts of blood recipients, an all‐ages and a younger subset, from a 2007 societal perspective. Model variable values were obtained from US screening data, Blood Systems Laboratory, the Health Care Utilization Project, and published literature. RESULTS: For the all‐ages cohort, compared to no testing, the cost‐effectiveness of testing all donors one time was $757,000 per QALY, all donors two times $970,000 per QALY, and universal testing $1.36 million per QALY. In the all‐ages and the younger transfused populations, testing donors with geographical exposure was most cost‐effective ($173,000 and $29,000/QALY, respectively). The most influential variables in the model were related to characteristics of the transfused population: survival and health state utilities. With respect to T. cruzi variables, results were most sensitive to seroprevalence and transmissibility. CONCLUSION: Selective T. cruzi screening generates nearly the same effectiveness as universal screening, but at a reduced cost. Outcomes and associated costs of Chagas disease take longer to materialize than the average life expectancy of transfusion recipients.