Analysis of reactive HCV results detected by current blood screening strategy / 中国输血杂志

【Objective】 To assess the status of HCV infection by analyzing the results of anti-HCV reactive blood samples detected by the current blood testing strategy, and discuss the viability of classified management of reactive blood donors. 【Methods】 The anti-HCV reactive samples (dual ELISA and once NAT), from May 2017 to October 2018, were divided into three groups: samples both anti-HCV and HCV RNA reactive, sole HCV RNA reactive, and sole anti-HCV reactive, and all of them were confirmed by recombinant immunoblot assay (RIBA). The positive predictive value (PPV) between groups were compared. The sensitivity, specificity and PPV for each reagent under different screening threshold (screening threshold for routine detection, optimal screening threshold, and corresponding screening threshold of the highest PPV) were analyzed. The group with low PPV were stratified by ELISA S/CO values, and PPV by different screening threshold was compared. 【Results】 There were 939 reactive samples (0.49%, 937/191 627). Confirmed by RIBA, the positive rate of anti-HCV reactive samples was 10.67%(100/937). Two samples were sole HCV RNA reactive (0.001%). Both anti-HCV+ HCV RNA reactive samples were 6.71%(63/939), with the PPV of 96.83%(61/63). Sole anti-HCV reactive samples were 93.08(874/939), with the PPV of 4.46%(39/874), among which PPV by dual and one ELISA reagent were 18.72% and 0.15%, respectively, showing statistically significant difference (P<0.05). The PPV between different S/CO values was statistically significant (P<0.05). The optimal screening thresholds of anti-HCV reagent were 9.29 and 3.97, according to the ROC curve, with significant difference noticed in PPV by different screening threshold (P<0.05). PPV in the sole anti-HCV reactive group increased from 4.46% (the routine screening threshold) to 49.35%(the optimal screening threshold), and the difference was statistically significant (P<0.05). 【Conclusion】 The blood donors with both anti-HCV and HCV RNA reactive can be determined as HCV infection and need to be permanently deferred. The S/CO value of sole anti-HCV reactive samples was positively correlated with RIBA confirmation results, and the higher the S/CO value, the greater the chances of positive confirmation are. With the current blood screening strategy, the HCV infection status of sole anti-HCV reactive blood donors can be determined by establishing a screening threshold with high PPV or adding confirmatory test.

The Role of the Signal-to-Cutoff Ratio in Automated Anti-HCV Chemiluminescent Immunoassays by Referring to the Nucleic Acid Amplification Test and the Recombinant Immunoblot Assay

BACKGROUND: Following discontinuation of the recombinant immunoblot assay (RIBA), the only available supplementary test for the detection of hepatitis C virus (HCV) is the nucleic acid amplification test (NAAT). However, the NAAT does not adequately detect past HCV. Consequently, it is hard to distinguish between past HCV infection and biological false positivity with an anti-HCV result alone. We assessed the diagnostic performance of two immunoassays: the ARCHITECT anti-HCV chemiluminescent microparticle immunoassay (CMIA; Abbott Diagnostics, Wiesbaden, Germany) and the Access HCV Ab PLUS chemiluminescent immunoassay (CIA; Bio-Rad, Marnes-la-Coquette, France). We also explored an optimized algorithm to determine the anti-HCV results. METHODS: We tested 126,919 patients and 44,556 individuals who underwent a medical checkup. RIBA and NAAT were conducted for samples that tested anti-HCV-positive using CMIA and CIA. We assessed the optimal signal-to-cutoff (S/CO) ratio in HCV-positive samples. RESULTS: In total, 1,035 blood samples tested anti-HCV-positive. Of these, RIBA was positive in 512, indeterminate in 160, and negative in 363 samples. One hundred sixty-five samples were NAAT-positive. Diagnostic sensitivity and positive predictive value (PPV) were 96.7% and 52.1%, respectively, for CMIA, and 94.7% and 72.3%, respectively, for CIA. The optimal S/CO ratio was 5.2 for CMIA and 2.6 for CIA at 95% PPV. In total, 286 samples tested positive in CMIA and 444 in CIA, while 443 samples tested positive in both assays. CONCLUSIONS: It is hard to determine anti-HCV positivity based on the S/CO ratio alone. However, this study elucidated the role of the S/CO ratio by using the NAAT and RIBA.

Analysis of anti-nuclear antibody and anti-nuclear antibody spectrum in 2 325 patients / 国际检验医学杂志

Objective To investigate the clinical value of anti-nuclear antibody(ANA) and anti-nuclear antibody spectrum(ANAs) detection.Methods A total of 2 325 patients with or suspected with autoimmune diseases(AID) were enrolled and detected for ANA and ANAs by using indirect immunofluorescence assay(IIF) and linear immunoblot assay(LIA) respectively.All detected results were analyzed.Results Among 2 325 patients,896 cases(38.54%) were positive with ANA,with positive rate of 45.46% in female patients,which was higher than the 18.46% of male patients(P<0.05),and the common fluorescence patterns were nuclear particle pattern,nuclear homogeneous pattern and the nucleolus pattern.816 cases(35.10%) were positive with ANAs,and the positive rates of anti-Sjogren's syndrome(SS)-B antibody,anti Ro-52 antibody and anti SS-A antibody were relatively higher.The consistency rate of the two methods was 91.66%.Conclusion ANA and ANAs detection could be with certain correlation,but might be not completely consistent,detection could improve the detection rate and reduce the missed detection rate.

Potential immunological markers for diagnosis and therapeutic assessment of toxocariasis / Estudo de marcadores imunológicos para o diagnóstico e avaliação terapêutica da toxocaríase

In human toxocariasis, there are few approaches using immunological markers for diagnosis and therapeutic assessment. An immunoblot (IB) assay using excretory-secretory Toxocara canis antigen was standardized for monitoring IgG, IgE and IgA antibodies in 27 children with toxocariasis (23 visceral, three mixed visceral and ocular, and one ocular form) for 22-116 months after chemotherapy. IB sensitivity was 100 percent for IgG antibodies to bands of molecular weight 29-38, 48-54, 95-116, 121-162, >205 kDa, 80.8 percent for IgE to 29-38, 48-54, 95-121, > 205 kDa, and 65.4 percent for IgA to 29-38, 48-54, 81-93 kDa. Candidates for diagnostic markers should be IgG antibodies to bands of low molecular weight (29-38 and 48-54 kDa). One group of patients presented the same antibody reactivity to all bands throughout the follow-up study; in the other group, antibodies decayed partially or completely to some or all bands, but these changes were not correlated with time after chemotherapy. Candidates for monitoring patients after chemotherapy may be IgG antibodies to > 205 kDa fractions, IgA to 29-38, 48-54, 81-93 kDa and IgE to 95-121 kDa. Further identification of antigen epitopes related to these markers will allow the development of sensitive and specific immunoassays for the diagnosis and therapeutic assessment of toxocariasis.

Métodos imunológicos desempenham papel importante no diagnóstico da toxocaríase, entretanto há poucos estudos sobre marcadores diagnósticos e de acompanhamento terapêutico. Foi padronizado ensaio de immunoblot (IB) empregando antígeno de excreção-secreção de Toxocara canis para pesquisa de anticorpos IgG, IgE e IgA em 27 crianças com toxocaríase nas formas visceral (23), mista visceral e ocular (3) e ocular (1), por 22-116 meses após quimioterapia. Foram observados dois perfis de reatividade dos anticorpos: permanência contra todas as frações no decorrer do estudo; diminuição ou negativação contra algumas ou todas as frações, porém, essas mudanças não se correlacionaram com tempo de tratamento. A sensibilidade do IB foi 100,0 por cento para anticorpos IgG específicos para frações de massa molecular de 29-38, 48-54, 95-116, 121-162, > 205 kDa, 80,8 por cento para IgE específicos para 29-38, 48-54, 95-121, > 205 kDa e 65,4 por cento para IgA específicos para 29-38, 48-54, 81-93 kDa. Anticorpos IgG específicos para frações de baixa MM (29-38 e 48-54 kDa) podem ser sugeridos como candidatos a marcadores diagnósticos. Por sua vez, anticorpos IgG para fração > 205 kDa, IgA para 29-38, 48-54, 81-93 kDa e IgE para 95-121 kDa podem ser candidatos a marcadores terapêuticos. A identificação de epítopos antigênicos relacionados a estes marcadores poderá ser importante para o desenvolvimento de ensaios altamente sensíveis e específicos no diagnóstico e avaliação terapêutica da toxocaríase.

Analysis of the Effectiveness of Blotting Assays for Donor Blood Screening / 대한수혈학회지

BACKGROUND: Immunoblot assays (IBAs) have been widely used to confirm the reactivity of immunoassay. However, indeterminate (ID) results have shown the limits for interpreting IBAs. There is some debate about the benefit of these assays. We assessed the actual status of the IBAs for the donor screening process and we proposed more available algorithms. METHODS: We analyzed the data from the blood information management system of the Korean Red Cross. This study was approved by the Institutional Review Board of the KRC. The analyzed data included the present condition of various utilities and the results of the IBAs in the world. RESULTS: The infectivity of the ID results in IBAs seemed not to be high, but the safety could not be assured. IBA for HTLV was used as a confirmatory test in many countries. Most of the eligible blood donors could be saved by IBAs. CONCLUSION: IBAs seem to be valuable methods as supplemental and follow up tests for ID results. Furthermore, IBAs were useful to distinguish eligible blood donors. When donors show positive results on an immunoassay and NAT (HIV and HCV) concurrently, then IBA does not seem to be required. Only a RIBA for HCV is recommended for the donors showing a S/CO ratio above 2.0 on immunoassay. The additional alternative immunoassay would be effective in the HTLV screening algorithm.

Comparison of enzyme-linked immunosorbent assay test with immunoblot assay in the diagnosis of pemphigus in Indian patients.

Background: The diagnosis of pemphigus vulgaris (PV) and pemphigus foliaceous (PF) rests upon clinical, histological and immunofluorescence features. Enzyme-linked immunosorbent assay (ELISA) test and immunoblot (IB) assay have shown variable sensitivity and specificity. Aims: We compared the utility of ELISA and IB in pemphigus patients. Methods: Sixty-six pemphigus cases (PV-54, PF-12) and 72 controls (other vesicobullous disorders and healthy controls) were inducted. ELISA for anti-Dsg 3 and 1 antibodies and IB assay were performed. Results: On ELISA, both mean anti-Dsg 1 and 3 titers were raised in PV and PF. Mean anti-Dsg 1 in mucocutaneous PV was significantly higher than in mucosal PV and mean anti-Dsg 3 was significantly raised in PV than in PF. Anti-Dsg 1 and 3 in the control group were negative. Sensitivity and specificity of ELISA in PV was 98.14% and 90.5% while in PF it was 91.6% and 61.1%, respectively.On IB in PV, 36 cases (66.67%) showed the 130 kDa and 160 kDa antigen bands, 12 (22.2%) only the 130 kDa and six (11.1%) only the 160 kDa band. Eight of the nine pure mucosal cases (88.8%) showed only the 130 kDa. In PF, only the 160 kDa antigen was detected. These antigens were not identified in the control group. Sensitivity and specificity of IB in PV was 88.9% and 100% and in PF it was 100% and 95.2%, respectively. Conclusion: Both tests could differentiate pemphigus from other dermatoses, including other blistering disorders. ELISA could not make a distinction between PV and PF or between the various clinical phenotypes of PV. IB differentiated between PV and PF and the different clinical variants of PV.

Evaluation of Anti-dsDNA Antibody Tests: Crithidia luciliae Immunofluorescence Test, Immunoblot, Enzyme-linked Immunosorbent Assay, Chemiluminescence Immunoassay / 대한진단검사의학회지

BACKGROUND: Anti-double stranded DNA antibody (anti-dsDNA) test is useful for the diagnosis and monitoring of systemic lupus erythematosus (SLE). Although several methods are available, none of them is completely satisfactory and differences among them have been reported. We evaluated the diagnostic performance of 6 commercial kits for anti-dsDNA detection. METHODS: A total of 142 sera (SLE [N=74], other systemic rheumatic diseases [N=50], other diseases [N=18]) were tested by 6 different assay kits using different antigenic sources of DNA: Crithidia luciliae immunofluorescence test (CLIFT), salmon testes (immunoblot, IB), human (ELISA I), salmon testes with nucleosome linker (ELISA II), plasmid (ELISA III), and synthetic oligonucleotides (chemiluminescence immunoassay, CLIA). RESULTS: With manufacturers' cut-off values, 6 test kits showed sensitivities of 55.4-91.9%. ELISA I had a greater sensitivity than the other five assays (P0.05). With cut-off values set at 95% of specificity, ELISA II had a higher sensitivity than ELISA III (63.5% vs. 41.9%, P<0.05). IB had poor concordance rates with other assays (42.0-65.0%). Pearson correlation coefficients among 4 quantitative assays were 0.667-0.798. CONCLUSIONS: Six different assays showed various performances depending on the methods and cut-off values used. Except IB, the other five assays can be used for the detection of anti-dsDNA.

Significance of antibody of liver antigens in diagnosis of autoimmune hepatic diseases / 中国基层医药

Objective To study the significance of anti-antibody of liver antigens in the diagnosis of autoim- mune hepatitis disease.Methods Patients were divided into three groups according the diseases:autoimmune hepatic disease 45 cases including autoimmune hepatitis(AIH)15 cases,primary biliary cirrhosis(PBC)20 cases,and prima- ry sclerotic cholangitis(PSC)10 cases;Various virus hepatitis 50 cases;Liver damage of unknown cause 30 cases.Au- to-antibody of liver antigens SLA/LP,LKM-1,LC-1,and AMA-M2 were identified by indirect immunofluorescence (IIF)assay and immunoblot assay.Results The positive rates of anti-SLA/LP(46.6%)was significantly higher than those of anti-LKM-1(13.3%),anti-LC-1(0%),and anti-AMA-M2(13.3%)in patients with AIR while these four antibodies were negative in patients with virus hepatitis.The positive rates of anti-AMA-M2 in patients of PBC and unknown liver damage were 95.0% and 6.6%,respectively.Conclusion Anti-SLA/LP is a new specific serum marker in diagnosis of AIR.The auto-antibody detection of liver antigens will be helpful to the diagnosis and therapy of autoimmune hepatic disease.

The Correlation between the S/CO Ratio of Anti-HCV ELISA, and the Results of the RIBA and RNA Test / 대한수혈학회지

BACKGROUND: Recombinant immunoblot assay (RIBA) or RNA test is considered to be a supplemental test for confirming a HCV infection. A correlation has been reported between the signal-to-cutoff (S/CO) ratios of a third generation HCV enzyme-linked immunosorbent assay (ELISA) and a confirmed HCV infection. This study examined the results of an evaluation of domestic anti-HCV EIA and immunoblot kit (RIBA) in Korean donors. METHODS: A total of 375,576 donor samples were tested for anti-HCV using the LG third generation HCV ELISA (LG HCD 3.0 TMB, LGphD, Korea) and HCV RNA by NAT (Biomerieux/Roche RT-PCR, 24 pool). The anti-HCV repeat reactive samples were further tested by third generation RIBA (LG HCD Confirm, LGphD, Korea). A positive result by either the nucleic acid amplification test (NAT) or RIBA was interpreted as a confirmed HCV infection. RESULTS: There were 506 out of the 375,576 donor samples (0.13%) that were anti-HCV repeat reactive (RR) by routine screening ELISA. The confirmed HCV prevalence in the donors was 0.01% (RIBA 42/375,570, RNA 36/375,570). 443 samples from the 506 repeat reactive samples in ELISA (87.6%) showed a S/CO ratio 3.6 (mean 4.40+/-0.80), compared with the negative group (mean 1.54+/-0.64). CONCLUSION: There was a good correlation between a high S/CO ratios and a confirmed HCV infection. In addition, samples showing a low S/CO ratio with an ID (Indeterminate) or negative RIBA result suggest a high probability of nonspecific reactivity in ELISA.

Qualitative and Quantitative Measurements of Anti-HCV Positive Blood Donor Group / 대한수혈학회지

BACKGROUND: To determine the positivity of hepatitis C virus-ribonucleic acid (HCV-RNA), we tested blood specimens of donor both positive in enzyme immunoassay (EIA) and in immunoblot assay, and those positive in EIA but indeterminate in immunoblot assay by nucleic acid amplification test (NAT). After quantifying HCV-RNA of specimens positive in NAT, we compared the titers of HCV-RNA between blood donor group and patient group. METHOD: One hundred twenty blood specimens positive both in screening test and in confirmative test, and 20 specimens positive in screening test but indeterminate were tested by qualifying NAT. After testing the specimens positive in this test by quantifying NAT, we classified specimens into 3 groups, normal group whose ALT values were within 45 IU/L, abnormal group whose values were higher than 45 IU/L and patient group who admitted into hospital to treat chronic hepatitis C and then compared HCV-RNA among groups. RESULTS: 81% of specimens both positive in screening test and in confirmative test was positive in NAT. Only 10% of specimens positive in screening test but indeterminate in confirmative test was positive in NAT. Ages of patient group were highest among groups and titers of HCV-RNA of patient group were lower than any other group. Correlation of AST/ALT values with the titers of HCV-RNA was not shown. CONCLUSION: It is concluded that the study groups show no difference of HCV-RNA titers whether they have symptoms of liver disease or not. The titer of HCV-RNA has no correlation with AST/ALT values.

Diagnostic Usefulness of Immunoblot Assay in Autoimmune Bullous Dermatoses / 대한면역학회지

Immunologic or immunopathologic assays are neccesary for the diagnosis of autoimmune bullous dermatoses including pemphigus vulgaris (PV), bullous pemphigoid (BP), and epidermolysis bullosa acquisita (EBA). The objectives of this study is to compare the sensitivity and usefulness of indirect immunofluorescence 0F) with that of immunoblot assay using amplified alkaline phosphatase staining system in the diagnosis of the above diseases; detection of disease-specific IgG autoantibodies. We selected 4 patients in each bullous dermatosis of PV, BP, and EBA, who had serum levels of IgG autoantibodies at a titer of 1:80 or higher. In each three disease, 2 patients with negative serum antibodies or serum titer lower than 1:20, were also enrolled. Among the former 4-patient groups the titers of IgG antibodies found on indirect IF were in the range of 1:80 to 1:160, whereas the titers recognized by immunoblot assay were 1 or 2 dilutions higher in most of these patients. In the latter 2-patient groups, 4 out of the 6 cases revealed antibody-positive on immunoblot-staining membrane. The indirect IF can be performed easily and seems favorable in the aspect of cost-effectiveness. However, immunoblot assay with sensitive staining method would be warranted in cases of antibody-negative or atypical clinical variants of autoimmunebullous dermatoses to confirm the diagnosis.

A Study on ALT levels in Hepatitis Marker Positive Donors -Anti-HBc Positive Rate in Blood Donors- / 대한수혈학회지

BACKGROUND: Hepatitis viral markers and ALT levels were evaluated in 190,679 blood donors to infer correlation between positive and control groups. The positive rate of anti-HBc was also observed in HBsAg and anti-HBs negative donors to know necessity about adding the item to the donor screening test. METHODS: The viral markers were tested by EIA method and ALT test was carried by auto-chemistry analyzer. Anti-HCV ELISA positive samples were confirmed by immunoblot assay. RESULTS: The rate of HBsAg(+) was 3.01% and that of anti-HCV was 0.54% of blood donors. The rate of HBsAg(+) and anti-HCV(+) was 0.01% of blood donors. Average ALT level of anti-HCV(+) (immunoblot assay) group was significantly higher than that of the control group. (p<0.001). The positive rate of anti-HCV in confirmatory test (immunoblot assay) was 15.76%. The proportion of donors who were HBsAg(-) and anti-HBs(-) and anti-HBc(+) was 1.87% in 1500 blood donors. CONCLUSION: The average ALT levels seem to be not correlated with viral hepatitis marker positivity except anti-HCV immunoblot assay positivity. It is suggested that anti-HBc and anti-HBs test should be added to screening test for donors and recipients to prevent post transfusional hepatitis.

A case: control study on the risk factors of hepatitis C virus infection among Koreans

In order to identify the risk factors for hepatitis C virus (HCV) infections, a case-control study was conducted from September 1993 to April 1994. HCV infection was confirmed by the second generation of recombinant immunoblot assay. Sixty-four cases and 128 controls matched for age and sex with a 1:2 ratio of cases to controls were enrolled. Exposure data were obtained from all participants by self-administered questionnaire and the odds ratios of possible risk factors of HCV infection analysed. Sixty-four cases consisted of forty-two patients with chronic hepatitis, nine with cirrhosis, one with hepatocellular carcinoma, and twelve with normal liver function. History of acute hepatitis (OR 3.9) and transfusion (OR 2.4) were associated with an increased risk of HCV infection. Operation, acupuncture, endoscopy, tooth extraction, tattooing, ear piercing, needle sharing and family history of hepatitis were not associated with an increased risk of HCV infection. In conclusion, transfusion remains the major route of transmission of HCV in Korea.

Analysis of auto-antibody profiles in 58 patients with PBC / 中国免疫学杂志

Objective: To study the role of auto-antibody in the diagnosis of patients with PBC. Methods: ANA, SMA and AMA in serum of 58 PBC patients were tested by indirect immune fluorescence and Western blot. Such auto-antibodies as Anti-type AMAM2,anti-SLA/LP, anti-LKM-1 and anti-LC-1 were also identified. Results: Auto-antibodies existing in patients with PBC were mainly AMA(96.5%) and AMAM2(93.1%) and the titer was beyond 1∶100. 8 cases of those patients were positive with ANA and SMA simultaneously. One case had positive AMA and SLA/LP in serum and the clinical appearances were the same as those of type Ⅰ and type Ⅲ autoimmune hepatitis. 19 patients with positive AMAM2 in serum had liver-puncture and the results suggested the diagnosis of PBC in 63.7%(12/9). Conclusion: Test of auto-antibodies is clinically significant for the diagnosis of autoimmune hepatitis.