ELISA检测抗-HCV灰区、弱反应标本与FQ-PCR检测结果对比分析

目的探讨丙型肝炎病毒抗体(抗-HCV)酶联免疫吸附试验(ELISA)检测呈灰区、弱阳性样本采用荧光定量PCR(FQ-PCR)的复检情况。方法选取2012年4月~2016年3月于本院手术和输血治疗的1 348例患者为研究对象,根据ELISA法抗-HCV检测结果将患者标本分为:A组:阴性(S/CO0.3)240例;B组:灰区(0.7≤S/CO1)902例;C组:弱反应性(1≤S/CO3.8)206例,所有研究对象均采用FQ-PCR复检,比较两种方法检测结果。结果 B组灰区和C组弱反应性标本中,FQ-PCR复检分别有41例(4.54%)和172例(83.49%)HCVRNA浓度1 000 IU/ml;ELISA检测抗-HCV双试剂灰区HCVRNAFQ-PCR阳性率高于单试剂灰区,差异有统计学意义(P0.05);ELISA检测抗-HCV双试剂弱反应性HCVRNA FQ-PCR阳性率与单试剂弱反应性比较,差异无统计学意义(P0.05)。结论 ELISA法检测抗-HCV为灰区、弱反应性标本存在一定的HCV RNA阳性标本漏检和误诊,采用FQ-PCR检测对于HCV感染早期诊断和治疗尤为重要。     

慢性丙型肝炎患者血清HCV-RNA水平与抗-HCV抗体和肝纤指标水平相关性分析

目的探讨丙型肝炎患者RNA水平与抗-HCV抗体和肝纤指标水平的关系。方法收集96例慢性丙型肝炎患者的血清,实时荧光定量PCR测定HCV-RNA和ELISA法检测抗-HCV抗体。化学发光法检测LN、PCⅢ、CⅣ及放免法检测HA。并对抗-HCV抗体、HCV-RNA水平与血清肝纤指标之间的关系进行分析。结果本研究的96例抗-HCV抗体阳性的慢性丙型肝炎患者中,HCV-RNA阳性52例,阳性率为54.2%。随着抗-HCV抗体的S/CO值升高,HCV-RNA检出率也在增高,分别为9.5%、37.1%和92.5%。高病毒载量组与低病毒载量组比较,血清HA、LN、PCⅢ及CⅣ水平的差异无统计学意义(P0.05)。结论慢性丙肝患者HCV-RNA阳性检出率与抗-HCV抗体的S/CO值有关,S/CO值越高,HCV-RNA阳性率越高。而RNA水平与肝纤指标水平没有相关性。     

静脉吸毒人群中血浆抗-HCVEIA法、WB法与HCVRNA荧光定量PCR法比较

目的了解吸毒人群中抗-HCV EIA法、WB法与HCV RNA检测分析的符合性,选取适合高危人群的HCV实验室诊断策略。方法使用2种EIA试剂进行筛查试验,初检阳性者用另一种抗体试剂复检和用FQ-PCR(荧光定量PCR)法检测HCV RNA,抗体复检阳性的样品用HCV WB检测试剂进行确认。结果血清学试验115份样本初检结果阳性108份,阴性7份;对初检阳性者进行复检,结果阳性90份,阴性18份。复检阳性者使用WB确认,89份阳性,1份不确定;EIA抗-HCV结果S/CO≥3.8有71份,与WB阳性符合率为98.6%。EIA复检阳性样本中HCV-RNA检出率为82.22%,复检阴性样本中有HCV RNA的检出率为77.78%(14/18)。结论抗-HCV并不与HCV RNA同时出现。吸毒人员HCV感染的检测,在EIA筛查基础上,对阴性样本检测HCV-RNA,阳性样本用WB试验进行确认,确保检测的准确性。     

乙型肝炎表面抗原检测结果分析及灰区设置探讨

目的通过对血液标本乙型肝炎表面抗原(HBsAg)检测反应性和弱反应性结果的分析,探讨酶联免疫吸附试验(ELISA)设置灰区的意义。方法采用高灵敏度进口和常规国产ELISA试剂对献血标本进行HBsAg血液筛查,比较两种试剂的检测结果 ;对0.7≤S/CO1灰区范围的弱反应性标本进行复检,探讨HBsAg检测灰区设置的范围及意义。结果共检测13965份无偿献血者标本,其中进口试剂检测HBsAg的阳性率为0.87%,国产试剂检测的阳性率为0.77%,进口试剂比国产试剂的检出阳性率高;对0.8≤S/CO1灰区范围的26例弱反应性标本进行再检,两种试剂合计有3份标本S/CO≥1,占11.5%。结论 HBsAg检测进口试剂比国产试剂的灵敏度高,有条件的血站在选择HBsAg检测试剂时应选择一遍进口试剂进行检测;HBsAg检测的灰区范围建议设定为域值(cut-off值)下的80%。     

国产抗丙型肝炎抗体检测试剂盒灰区范围的设置

目的设置上海地区临床实验室常用的4种国产抗丙型肝炎病毒(HCV)抗体(简称抗HCV)检测试剂的灰区范围。方法收集抗HCV临床初检阳性样本656例,分别用荣盛、新波、科华和科美4家国产抗HCV检测试剂进行复检,分别计算初检不同S/CO值范围的复检率;选择抗HCV初、复检结果不一致的弱阳性样本338例,采用确认试验[重组免疫印迹法(RIBA)]进行确认,对RIBA结果为不确定的样本采用实时荧光定量聚合酶链反应(PCR)检测HCV RNA。以RIBA和HCV RNA结果作为金标准,绘制受试者工作特征(ROC)曲线,探讨其最佳临界S/CO值。以95%真阳性和95%真阴性S/CO值确定4种国产抗HCV检测试剂阴、阳性灰区范围。结果 4种国产抗HCV检测试剂的阴性复检符合率均90%,S/CO值均12.01;阳性复检符合率均95%。荣盛、新波、科华及科美4种国产试剂检测抗HCV的最佳临界S/CO值分别为1.31、2.48、3.22和4.32。分别以95%真阳性率和95%真阴性率确定4种国产抗HCV检测试剂的阳性和阴性临界值,得出荣盛、新波、科华及科美4种国产抗HCV检测试剂S/CO值的灰区范围分别为0.6~1.3、0.7~2.5、0.7~4.0和0.7~4.3。结论确认了上海地区4种常用国产抗HCV检测试剂的最佳临界S/CO值,为临床提供了4种抗HCV检测试剂的灰区范围。     

HCV-RNA、抗-HCV、肝功能指标对丙型肝炎的临床诊断价值

目的分析丙型肝炎病毒抗体(抗-HCV)阳性患者丙型肝炎病毒核酸RNA(HCV-RNA)载量、肝功能指标在丙型肝炎诊断中的价值。方法选取893例抗-HCV1 S/CO的患者,检测其HCV-RNA和丙氨酸氨基转移酶(ALT)、天门冬氨酸氨基转移酶(AST)、γ-谷氨酰基转肽酶(GGT)及α-L-岩藻苷酶(AFU)水平。采用受试者工作特征曲线(ROC曲线)分析各项指标诊断丙型肝炎的效能。结果抗-HCV、ALT、AST诊断丙型肝炎的ROC曲线下面积分别为0.884、0.760、0.760。抗-HCV截断值为10.26 S/CO时,其诊断丙型肝炎的灵敏度为95.7%,特异度为75.6%;ALT的灵敏度和特异度分别为68.6%、73.3%,AST的灵敏度和特异度分别为77.7%、65.3%。抗-HCV≥10.26 S/CO组HCV-RNA、肝功能指标均高于抗-HCV10.26 S/CO组,差异均有统计学意义(P0.05)。HCV-RNA阳性组抗-HCV、肝功能指标均高于HCV-RNA阴性组,差异均有统计学意义(P0.05)。抗-HCV与HCV-RNA载量等级的相关性最高(r=0.632,P0.05)。结论抗-HCV≥10.26 S/CO时,结合HCV-RNA、ALT、AST检测对丙型肝炎具有较准确和全面的诊断价值。     

献血者HBsAg、抗-HCV、抗-HIV、抗-TPELISA检测阳性与确证试验的对比研究

目的评价ELISA两步法用于献血者血液HBsAg、抗-HCV、抗-HIV、抗-TP筛检阳性标本与确证试验结果的符合性,探讨确证试验结果用于献血者管理、酶免试剂选择和ELISA灰区范围设置的理论依据。方法 HB-sAg采用电化学发光(ECLIA)中和试验、抗-HCV采用重组免疫印迹试验(RIBA)、抗-HIV采用蛋白质印迹(WB)法、抗-TP采用凝集法(TPPA)确证试验对ELISA两步法检测阳性及灰区标本456份进行检测,将2者结果对比分析。结果 HBsAg、抗-HCV、抗-HIV和抗-TP确认阳性率分别为41.18%、38.00%、18.52%和48.82%;抗-HIV、抗-TP灰区(S/CO值0.5～1.0)标本确证试验无阳性,HBsAg灰区(S/CO值0.8～1.0)、抗-HCV灰区(S/CO值0.7～1.0)均有阳性和不确定标本检出。结论为ELISA两步法应用于献血者酶免4项检验效果进行了科学评价,为假阳性献血者检验结果的告知及ELISA灰区设立原则提供了实验依据。     

天津地区无偿献血者HCVRNA与抗-HCV、ALT检测结果相关性分析

目的探讨本中心无偿献血者丙型肝炎病毒RNA(HCV RNA),与丙型肝炎抗体(抗-HCV)和丙氨酸氨基转移酶(ALT)检测结果之间的相关性。方法经ELISA检测抗-HCV阳性标本313例,采用转录介导的扩增(TMA)-化学发光法定性检测HCV RNA,速率法检测ALT水平。结果 313例抗-HCV阳性血清中HCV RNA阳性者141例(阳性率45.05%);抗-HCV S/CO值1-3.79实验组,HCV RNA阳性率为5%,抗-HCV S/CO值为3.80-4.99的实验组,HCV RNA阳性率为76.74%,S/CO值≥5时,HCV RNA阳性率为56.25%。各组之间差异具统计学意义(P<0.05);抗-HCV(+)/HCV RNA(-)组与抗-HCV(+)/HCV RNA(+)组的ALT检测异常率,分别为17.44%和17.73%,2组之间差异无统计学意义。结论 HCV RNA阳性率与抗-HCV的S/CO值有一定相关性;在抗-HCV异常的无偿献血者人群中,ALT异常率与HCV RNA检测结果无相关性。     

维持血液透析患者丙型肝炎抗-HCV ELISA检测灰区标本确认的意义

目的探讨维持血液透析患者酶联免疫吸附试验(ELISA)抗-HCV检测灰区标本确认的临床价值。方法选取ELISA检测抗-HCV吸光度/临界值(S/CO)比值在0.5~3.5之间的标本95份用荧光定量聚合酶链反应(PR-PCR)进行确认。结果在抗-HCV灰区(0.5≤S/CO3.5)的95份标本中,0.5≤S/CO1.0的标本13份,经PR-PCR荧光定量进行确认阳性2份,阳性率15.4%(2/13);1.0≤S/CO3.5两个区段标本82份,经PR-PCR确认25份,阳性率分别33.3%(10/30)和28.8%(15/52)。结论维持血液透析患者(MHD)抗-HCV ELISA检测灰区标本应进一步确认,以防止漏诊误诊。     

ORTHO抗-HCV酶联免疫试剂2种孵育试验程序在血液筛查中的比较研究

目的确定1种3代抗-HCV酶免试剂(ORTHO HCV 3.0 ELISA试剂)2种孵育试验过程在血液筛查中是否存在差异。方法随机留取常规献血者血液筛查中检出的抗-HCV反应性的血液标本180(人)份作HCVRNA检测,并用RIBA和不同于血站常规抗-HCV筛查的酶免试剂作抗-HCV复测;依据ORTHO HCV 3.0 ELISA检测试剂说明书中的长、短2种孵育检测程序,同时作对比检测。结果 180份初筛抗-HCV阳性标本中,有16份ORTHO HCV 3.0 ELISA 2种检测程序的检测结果不一致,其中5份为短孵育试验反应性、11份为长孵育试验反应性,短孵育试验漏检2份被确证抗-HCV阳性标本。ORTHO HCV 3.0 ELISA试剂的长孵育检测程序灵敏度高于短孵育试验程序,2种试验程序的S/CO值分布没有差别,但RIBA不确定标本其S/CO值分布在一定的灰区范围内。结论在献血人群血液筛查中,采用具有更高灵敏度的长孵育酶免程序和设定合理的结果判定灰区,有助于预防输血传播HCV,提高血液的安全。     

丙型肝炎病毒抗体联合核酸定量检测对丙型肝炎患者的早期诊断价值研究

目的：探讨在丙型肝炎患者早期诊断中丙型肝炎病毒抗体联合丙型肝炎病毒核酸（HCV RNA）检测的临床价值。方法对116例鼓楼医院住院及门诊丙型肝炎患者采用实时荧光定量聚合酶链反应（FQ-PCR）测定HCV RNA、酶联免疫吸附试验（ELISA）和金标法测 HCV抗体、速率法测丙氨酸氨基转移酶（ALT ）。结果116例丙型肝炎患者中,ELISA HCV抗体阳性97例（占83．6％）,HCV RNA阳性85例（占73．3％）,HCV 金标阳性77例（占66．4％）,ALT阳性69例（占59．5％）,HCV抗体和 HCV RNA联合检测总阳性率（指任-指标阳性即为阳性）为100．0％。结论 HCV抗体和HCV RNA联合检测扩大了丙型肝炎检测范围,降低了丙型肝炎的漏诊率,有利于丙型肝炎的早期诊断。     

Significance of the signal-to-cutoff ratios of anti-hepatitis C virus enzyme immunoassays in screening of Chinese blood donors

BACKGROUND: The correlation between signal-to-cutoff (S/CO) ratios of a second-generation hepatitis C virus (HCV) enzyme immunoassay (EIA; Abbott) and a third-generation HCV enzyme-linked immunosorbent assay (ELISA; Ortho) and confirmed HCV infection has been reported. The utility of the values for the Chinese anti-HCV EIA kits, however, has not been studied in evaluating test results in Chinese blood donors. STUDY DESIGN AND METHODS: A total of 156 donor samples repeat reactive for anti-HCV at routine screening from five representative regions of China were retested for anti-HCV by the Ortho third-generation HCV ELISA and six Chinese EIA kits and for HCV RNA by a human immunodeficiency virus-1 and HCV assay (Procleix, Chiron Corp.). The HCV RNA-nonreactive samples were further tested for anti-HCV by a third-generation recombinant immunoblot assay RIBA (Chiron Corp.). The positive result by either nucleic acid amplification test or RIBA was interpreted as confirmed HCV infection. RESULTS: The confirmed HCV prevalence rate in donors in five representative regions obtained in this study was 0.20 percent (77/37,900) in 2004. All seven anti-HCV EIA kits had a significant correlation between S/CO ratios and confirmed HCV infection. The threshold S/CO ratios, which predicted more than 95 percent of confirmed HCV infections for the Ortho, SABC, BGI-GBI, InTec, GWK, KHB, and WANTAI kits, were 3.8, 6.0, 7.0, 8.6, 10.0, 10.0, and 14.0, respectively. CONCLUSIONS: Anti-HCV EIA kits commonly used in Chinese donors screening demonstrate good correlation between S/CO ratios and the confirmed infection. For the Ortho third-generation HCV ELISA, the S/CO ratio of 3.8 determined by the US Centers for Disease Control and Prevention is applicable to Chinese blood donors. The Chinese domestic EIA kits evaluated show a diverse range of threshold S/CO ratios.     

探讨丙肝核心抗原检测在母婴传播中的应用价值

目的：探讨丙肝核心抗原检测在母婴传播中的应用价值。方法选取选取2014年1月～2015年1月在我院就诊的HCV‐Ab（ELISA）阳性产妇100例作为研究对象,产妇于生产前采集静脉血3～5 mL ,新生儿采集脐血2～3 mL 进行检测。首先使用HCV‐RNA 对100例HCV‐Ab（IGg）阳性产妇血清标本进行检测。使用HCV‐cAg、HCV‐Ab、HCVRNA三种方法对新生儿脐血标本进行检测,观察三种方法的阳性率。结果 HCVcAg阳性率与 HCV‐Ab（IgG）的阳性率比较差异具有统计学意义,P＜0．05。HCVcAg阳性率与HCV‐RNA阳性率比较,差异没有统计学意,P＞0．05。结论 HCVcAg检测在HCV母婴传播初期诊断中,可作为临床诊治的依据,值得基层医院推广。     

丙型肝炎抗原检测

目的探讨在HCV感染早期,HCV Ab为阴性时用酶联免疫法检测HCV核心抗原的试验方法的可行性。方法用重组HCV核心抗原,免疫小鼠制备单克隆抗体,对HCV核心抗原进行酶联免疫法检测。结果HCV核心抗原检测灵敏度高可达5ng/ml,用酶联法对11份HCV Ab阴性,HCV RNA阳性标本检测9份阳性。结论酶联免疫法检测HCV核心抗原可作为HCV感染早期的抗原检测方法,较HCV RNA检测法简便、快速,可以作为核酸检测法的一种简易替代方法。     

The significance of repeat testing in Turkish blood donors screened with HBV,HCV and HIV immunoassays and the importance of S/CO ratios in the interpretation of HCV/HIV screening test results and as a determinant for further confirmatory testing

The purpose of this study was to investigate the intra‐assay correlations amongst initial reactive and repeat screening results used in enzyme immunoassays (EIAs) for hepatitis B virus (HBV), hepatitis C virus (HCV) and HIV in blood donors. This study evaluated the value of using the power of the signal to cut‐off (S/CO) ratio index for confirming anti‐HCV/HIV reactive screening results, thereby touching upon the utility of S/CO indices in determining whether further confirmatory testing was necessary. Screening test results of the 72 695 blood donors were evaluated over a 1‐year period. Correlation analysis among each initial test and retests was done by Pearson r test. Appropriate S/CO values to determine the need of the confirmation testing was investigated by ROC analyses. EIA intra‐assay correlations were of statistical significance and were determined as follows: 0·948 for anti‐HCV, 0·827 for anti‐HIV and 0·948 for HBsAg. The threshold S/CO ratio values which predicted more than 95% of the confirmation test result were 3·8 for HCV and 5·6 for HIV. We were able to demonstrate a strong level of intra‐assay correlation amongst EIAs, thereby eliminating the need for repetition of the screening test. Hence, we suggest that repeat screening should only be limited to HBV and HIV tests with low EIA S/CO ratios. Thus, using the power of the S/CO ratio in determining the need for HCV confirmation testing can be a cost‐effective measure, especially if the S/CO value is ≥3·8.     

High antibody level: an accurate serologic marker of viremia in asymptomatic people with hepatitis C infection

BACKGROUND: The screening and diagnosis of hepatitis C virus (HCV) infection is initiated by testing for antibody to HCV (anti‐HCV). A positive anti‐HCV test in blood donors represents ongoing infection in only a variable proportion of individuals. Because a high anti‐HCV level has been associated with viremia, a study was conducted to determine whether a high antibody level is an accurate serologic marker for viremia in asymptomatic anti‐HCV–positive persons. STUDY DESIGN AND METHODS: In a diagnostic test study, we included 856 anti‐HCV–positive blood donors in a blood bank at Guadalajara, Jalisco, Mexico, between 2002 and 2007. A third‐generation amplified chemiluminescence assay (ChLIA HCV) was used to detect anti‐HCV. A positive result of the qualitative nucleic acid test (HCV RNA) was considered the gold standard for viremia. RESULTS: By receiver operating characteristic analysis, the signal‐to‐cutoff (S/CO) ratio of 20 or more was chosen as optimal to identify viremia and so was defined as high anti‐HCV level. There was a significant difference in the proportion of viremia between subjects with high antibody level and those with lower levels (93.7% vs. 1.8%, respectively; p < 0.001). A high antibody level showed a sensitivity for viremia of 96.6% (95% confidence interval [CI], 93.8%‐98.1%), a specificity of 96.6 % (95% CI, 94.8%‐97.8%), and a likelihood ratio of 28.6 (95% CI, 18.4%‐44.6%). CONCLUSION: A high antibody level (S/CO ratio ≥20 by ChLIA HCV) clearly divides the viremic from the nonviremic blood donors and functions as an accurate serologic marker to guide the use of routine HCV RNA testing to confirm hepatitis C infection.     

HBsAg酶联免疫吸附试验灰区设置研究

目的评价与验证该实验室乙型肝炎表面抗原(HBsAg)试验设置0.9倍临界值(CO值)的合理性。方法参照美国临床和实验室标准协会(CLSI)发布的EP12-A2指南,通过试验确定HBsAg试验C5~C95区间即灰区。对HBsAg灰区标本进行抗体确认试验。绘制受试者工作特征曲线(ROC曲线)确定HBsAg试验最佳CO值。通过实验室既往数据,分析乙型肝炎病毒DNA(HBV-DNA)单阳性标本酶联免疫吸附试验(ELISA)结果分布(S/CO值)与灰区的关系。结果 (C50±20%)水平检测结果阴性数和阳性数均大于或等于95%,(C50-20%)~(C50+20%)水平范围包含C5~C95区间,灰区范围应在0.712~1.103倍CO值区间内。对44例HBsAg灰区标本(S/CO值0.900~0.990)进行中和试验,结果均为无反应性。绘制HBsAg的ROC曲线,曲线下面积(AUC)为0.981,最适CO值为0.063(现用CO值在0.055~0.060)。2010年11月2日至2013年12月31日检测标本研究886 291例患者,HBsAg阳性标本包括135例灰区标本,其中7例核酸检测法(NAT)检测结果均为反应性;共检出HBV-DNA单阳性标本421例,其HBsAg检测结果(S/CO值)分布区间为0.200~0.400,与阴性标本分布区间重叠、距0.9倍CO值较远。结论该实验室现阶段HBsAg试验设置0.900的CO灰区过于严苛,试验结果支持取消灰区设置。报道所提供的4种灰区评价方法为其他实验室在设置ELISA试验灰区方面提供了1种思路。     

应用ROC曲线分析确定ELISA检测抗-HBe灰区

目的应用受试者工作特征曲线（ROC曲线）分析方法确定酶联免疫吸附试验（ELISA）检测乙型肝炎e抗体（抗-HBe）的灰区,为临床判读提供合理的指导。方法 ELISA检测血清抗-HBe,留取S/CO介于0.5～1.7的标本,用美国雅培Axsym化学发光仪对其复检,结合临床症状及乙型肝炎病毒标志物定量结果,筛选结果可靠的标本,进行ROC曲线分析,根据ROC曲线分别确定其灰区的上、下限。结果根据ROC曲线分析法计算得出检测灰区为0.827～1.576。结论 ROC曲线法是设定ELISA检测灰区的较理想方法,非灰区部分具备较低的假阳性率及假阴性率,能增加不同医院间结果的可比性,减少不必要的医疗纠纷。