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1.  Serial follow-up of repeat voluntary blood donors reactive for anti-HCV ELISA 
Background:
Voluntary non-remunerated repeat blood donors are perceived to be safer than the first time blood donors. This study was planned for follow-up of previous hepatitis C virus (HCV) test results of anti-HCV enzyme-linked immunosorbent assay (ELISA) reactive repeat blood donors. The aim was to suggest a protocol for re-entry of the blood donors who are confirmed HCV negative by nucleic acid test (NAT) and recombinant immunoblot assay (RIBA). A group of repeat voluntary donors were followed retrospectively who became reactive on a cross sectional study and showed HCV reactivity while donating blood regularly.
Material and Methods:
A total of 51,023 voluntary non remunerated blood donors were screened for anti-HCV ELISA routinely. If anybody showed positivity, they were tested by two ELISA kits (screening and confirmatory) and then confirmed infection status by NAT and or RIBA. The previous HCV test results of repeat donors reactive by anti-HCV ELISA were looked back from the records. Data of donors who were repeat reactive with single ELISA kit (in the present study) were analyzed separately from those reactive with two ELISA kits (in the present study).
Results:
In this study, 140 (0.27%) donors who were reactive by anti HCV ELISA were included. Out of them, 35 were repeat voluntary donors and 16 (11.43%) were reactive with single ELISA kit. All 16 donors were reactive by single ELISA kit occasionally in previous donations. Their present ELISA positive donations were negative for HCV NAT and RIBA. A total of 19 (13.57%) donors were reactive with two ELISA kits. In their previous donations, the donors who were reactive even once with two ELISA kits were consistently reactive by the same two ELISA kits in their next donations also.
Conclusion:
Donor sample reactive by only single ELISA kit may not be considered as infectious for disposal as they were negative by NAT and or RIBA. One time ELISA positivity was found probably due to ELISA kit specificity and sensitivity. Donors reactive with two ELISA kit should be discarded as there is a high positivity with NAT/ RIBA. However, donors reactive by two ELISA kits and negative by NAT and RIBA should be followed up and may not be deferred permanently.
doi:10.4103/0973-6247.75979
PMCID: PMC3082711  PMID: 21572711
Anti-HCV ELISA; repeat voluntary blood donor; occult infections; donor follow-up; nucleic acid test; recombinant immunoblot assay
2.  Half a decade of mini-pool nucleic acid testing: Cost-effective way for improving blood safety in India 
Background and Objectives:
It is well established that Nucleic acid testing (NAT) reduces window phase of transfusion transmissible infections (TTI) and helps improve blood safety. NAT testing can be done individually or in pools. The objectives of this study were to determine the utility, feasibility and cost effectiveness of an in-house minipool-NAT(MP-NAT).
Materials and Methods:
Blood donors were screened by history, tested by ELISA and sero-negative samples were subjected to an in-house NAT by using reverse transcriptase-polymerase chain reaction (RT-PCR). Testing was done in mini-pools of size eight (8). Positive pools were repeated with individual samples.
Results:
During the study period of Oct 2005-Sept 2010 (5 years) all blood donors (n=53729) were screened by ELISA. Of which 469 (0.87%) were positive for HIV-1, HBV or HCV. Sero-negative samples (n=53260) were screened by in-house MP-NAT. HIV-NAT yield was 1/53260 (n=1) and HBV NAT yield (n=2) was 1/26630.
Conclusion:
NAT yield was lower than other India studies possibly due to the lower sero-reactivity amongst our donors. Nevertheless it intercepted 9 lives including the components prepared. The in-house assay met our objective of improving blood safety at nominal cost and showed that it is feasible to set up small molecular biology units in medium-large sized blood banks and deliver blood within 24-48 hours. The utility of NAT (NAT yield) will vary based on the donor population, the type of serological test used, the nature of kit employed and the sensitivity of NAT test used. The limitations of our in-house MP-NAT consisted of stringent sample preparation requirements, with labor and time involved. The benefits of our MP-NAT were that it acted as a second level of check for ELISA tests, was relatively inexpensive compared to ID-NAT and did not need sophisticated equipment.
doi:10.4103/0973-6247.126688
PMCID: PMC3943143  PMID: 24678172
Blood donor testing; blood safety; in house assay; mini-pool nucleic acid testing; real-time-polymerase chain reaction
3.  Prospective study of the meaning of indeterminate results of the Recombinant Immunoblot Assay for hepatitis C virus in blood donors 
Blood Transfusion  2008;6(2):107-111.
Background
The interpretation of “indeterminate” results of the recombinant immunoblot assay (RIBA) is a particularly sensitive issue for Transfusion Services, and donors with such a serological condition require long-term follow-up.
Materials and methods
In the Immunohaematology and Transfusion Medicine Division of Umberto I University Hospital (Rome, Italy), 102,979 donor blood units were screened for hepatitis C virus (HCV) antibodies by enzyme-linked immunosorbent assay (ELISA) over a 5-year period (01.01.2000 – 31.12.2004). Since 24.10.2001, HCV -RNA testing was added. All samples repeatedly reactive by ELISA were then submitted to a HCV confirmatory assay (RIBA).
Results
Among the 102,979 donors we found 271 positive to HCV ELISA testing. The results of the RIBA assay for these donors were negative in 178 (65.7%) cases, positive in 28 (10.3%) and indeterminate in 65 (24.0 %).
Of the 65 subjects with an indeterminate pattern, 24 completed a sufficient follow-up (median 25 months; range, 6 – 52), during which some (n=8; 33%) converted to a negative status, some (n=16; 67%) maintained their reactivity pattern, but none became seropositive for HCV.
Conclusions
The HCV-RIBA indeterminate status may indicate either a non-specific reaction (false positive) or a real pre-existing or initial infection and does not, therefore, enable a prediction of outcome. The use of HCV genomic assays (nucleic acid amplification testing), which are more specific than antibody-based assays (ELISA, RIBA), therefore improves HCV blood donor testing by allowing an accurate interpretation of such primary assays.
doi:10.2450/2008.0037-07
PMCID: PMC2626846  PMID: 18946955
hepatitis C virus (HCV); HCV-RIBA indeterminate; blood donors
4.  Hepatitis C virus in blood samples from volunteer donors. 
Journal of Clinical Microbiology  1993;31(3):606-609.
The specificities of four assays for hepatitis C virus (HCV) were compared by using units from volunteer blood donors. Upon Food and Drug Administration licensure of the first immunoassay for anti-HCV, EIA-1, units previously deemed acceptable for transfusion and all subsequent blood donations were screened. EIA-1 repeat-reactive (RR) units were tested for HCV by a second-generation enzyme-linked immunoassay (EIA-2) and by a four-antigen recombinant immunoblot assay (RIBA II) and for HCV RNA by reverse transcriptase polymerase chain reaction. All HCV RNA-positive samples were reactive by both RIBA II and EIA-2. All RIBA II-reactive sera were EIA-2 RR. In EIA-1, 0.45% of the prescreened units and 0.59% of the prospectively screened donors were RR. Of these, 52.5 and 54%, respectively, were EIA-2 RR, 71.4 and 69% of the EIA-2 RR units were reactive on RIBA II, and 93 and 88% of the RIBA II-reactive samples were HCV RNA positive. When the sample/cutoff ratio for EIA-2 was greater than 5, 91% of the samples were RIBA II reactive and 82% of the samples were HCV RNA positive. None of EIA-2 RR units with a sample/cutoff ratio of < 5 was RIBA II reactive or HCV RNA positive. In conclusion, RIBA II and RT PCR results are highly concordant. A sample/cutoff ratio of > 5 in EIA-2 is a good discriminator for the likelihood of a true HCV infection on the basis of RT PCR and RIBA II assays.
PMCID: PMC262828  PMID: 8384627
5.  Laboratory diagnosis of viral hepatitis C 
Objectives:
A retrospective study was carried out to assess the performance of hepatitis C diagnostic assays in our laboratory, and to determine the prevalence of hepatitis C among blood donors at the Sultan Qaboos University Hospital.
Methods:
From 1991 to 2001, approximately 55,000 serum samples collected from blood donors and patients were submitted to our laboratory for testing. All sera were screened for antibodies to hepatitis C virus (HCV) by three successive generations of the enzyme-linked immunosorbent assay (ELISA). Anti-HCV positive sera were further tested by recombinant immunoblot assay (RIBA). Reverse-transcriptase polymerase chain reaction (RT-PCR) for HCV RNA was carried out on a limited number (241) of ELISA positive samples.
Results:
Out of 30012 samples from blood donors that were screened for anti-HCV, 272 (0.91%) were positive. Of these, 46.5% were confirmed positive by RIBA. The proportion of patient sera that were confirmed positive varied from 95% among intravenous drug users to 81% in patients with hepatitis to 70% in those with haemoglobinopathies. HCV RNA was detected in 67%, 6%, and 0% of the RIBA positive, indeterminate and negative samples respectively.
Conclusions:
Based on RIBA, the prevalence of anti-HCV among blood donors in Oman is close to 0.5%. In our experience, RIBA-positivity is predictive of HCV infection in two thirds of subjects, and HCV infection is highly unlikely in those who are RIBA-negative. The experience at SQUH with three types of HCV assays has enabled the laboratory to develop a test algorithm, starting with screening anti-HCV ELISA.
PMCID: PMC3174725  PMID: 24019730
hepatitis C virus; ELISA; RIBA; polymerase chain reaction
6.  The Clinical Relevance of Persistent RIBA Indeterminate Reactions: Insights into the Natural History of HCV infection and Implications for Donor Counseling 
Transfusion  2012;52(9):1940-1948.
BACKGROUND
A solid phase recombinant-immunoblot-assay(RIBA) is often used to determine the specificity of antibody to hepatitis-C-virus(anti-HCV). The RIBA result is recorded as positive, negative or indeterminate. The interpretation of RIBA indeterminate reactivity and its significance to patients and blood donors are unclear. We attempted to address the clinical relevance of RIBA-indeterminate reactions in the context of the natural history of HCV infection in a prospectively followed cohort of anti-HCV positive blood donors.
STUDY-DESIGN AND METHODS
Donor demographics, HCV exposure history, humoral and cell-mediated immunity(CMI) to HCV were compared in 15 RIBA-indeterminates, 9 chronic-HCV-carriers and 8 spontaneously-recovered subjects. Serum samples were tested for the presence of anti-HCV by a liquid phase Luciferase-Immunoprecipitation-System(LIPS) assay. CMI was assessed by IFN-γ-ELISpot assay.
RESULTS
In the quantitative LIPS assay, the sum of antibody responses to 6 HCV-antigens showed significant (p<0.001) step-wise diminution progressing downward from chronic-carriers to spontaneously-recovered to RIBA-indeterminates. CMI responses in RIBA-indeterminates were similar to spontaneously-recovered subjects, and greater than chronic-carriers and negative controls (p<0.008). A parenteral risk factor was identified in 13% of RIBA-indeterminates as compared with 89% of chronic-carriers and 87% of spontaneously-recovered subjects. On average, donors in the RIBA-indeterminate group were older than the other groups.
CONCLUSION
The combined CMI and LIPS results suggest that persistent RIBA-indeterminate reactions generally represent waning anti-HCV responses in persons who have recovered from a remote HCV infection. In such cases, detectable antibody may ultimately disappear leaving no residual serologic evidence of prior HCV infection, as previously reported in a minority of long-term HCV-recovered subjects.
doi:10.1111/j.1537-2995.2011.03524.x
PMCID: PMC3346857  PMID: 22304422
HCV; RIBA indeterminate; HCV infection spontaneously recovered; Chronic HCV infection; RIBA 3.0; Cell-mediated immunity; IFNγ; Luciferase immunoprecipitation system (LIPS) assay
7.  Detection of hepatitis C virus by PCR in second-generation enzyme immunoassay-seropositive blood donors by using matched pairs of fresh frozen plasma and pilot tube sera. 
Journal of Clinical Microbiology  1996;34(9):2191-2195.
Between April 1993 and March 1995, 429 of 334,454 (0.13%) blood donations at the Toronto Centre of the Canadian Red Cross were reactive for hepatitis C virus (HCV) by second-generation enzyme immunoassay (EIA-2). Of the 429 EIA-2-positive donations, 189 (44%), 138 (32%), and 102 (24%) were positive, indeterminate, and negative by Second-Generation Recombinant Immunoblot Assay (RIBA-2). To assess HCV viremia and minimize the risk that specimen handling affected PCR-based detection, the qualitative AMPLICOR HCV test was performed on both pilot tube sera (PTS) and the corresponding fresh frozen plasma (FFP) from 294 EIA-2-reactive donations. AMPLICOR PCR results for PTS and FFP were 100% concordant and were confirmed by nested HCV PCR for 27 of 294 donations. The AMPLICOR HCV test was positive for 127 of 140 (91%) of RIBA-2-positive donations (81, 91, and 96% of donations with two, three, and four reactive bands, respectively), 5 of 88 (5.7%) indeterminate donations, and 0 of 66 (0%) RIBA-2-negative donations. The Third-Generation Recombinant Immunoblot Assay (RIBA-3) was performed on RIBA-2-negative, -indeterminate, and -positive, PCR-negative donations. RIBA-3 demonstrated enhanced specificity and resolved 18 of 88 (20%) of RIBA-2-indeterminate samples as HCV antibody positive. The study demonstrates that PTS are as suitable as FFP for PCR-based detection of HCV and can be used to determine if EIA-2-reactive blood donors are viremic at the time of donation.
PMCID: PMC229215  PMID: 8862583
8.  Significance of indeterminate third-generation hepatitis C virus recombinant immunoblot assay. 
Indeterminate hepatitis C virus (HCV) third-generation recombinant immunoblot assay (RIBA3.0; Ortho Diagnostic Systems) patterns were arbitrarily defined by the manufacturer as the detection of only one antibody out of the four that were sought, namely, c100 (NS4 encoded), c22 (core encoded), c33c (NS3 encoded), and NS5 (NS5 encoded). The aims of the present study were (i) to determine the prevalence of indeterminate RIBA3.0 patterns in patients consecutively tested for anti-HCV antibodies in a university hospital; (ii) to evaluate the significance of these patterns in terms of viral replication, liver disease, and risk factors for HCV; and (iii) to get an insight into the mechanism underlying this peculiar immune response. Among 3,074 serum samples consecutively tested for anti-HCV antibodies, 588 were found to be positive by screening assays. Fifty-nine of them (10%) were RIBA3.0 indeterminate and were compared with 59 RIBA3.0-positive ones. Thirty-one RIBA3.0-indeterminate and 53 RIBA3.0-positive serum samples were HCV RNA positive by PCR (53 versus 90%; P < 10(-6). RIBA3.0-indeterminate and RIBA-3.0-positive patients with positive PCR results were not significantly different for the prevalence of risk factors for HCV infection and elevated serum alanine aminotransferase activities. Immunosuppression, attributable to coexisting human immunodeficiency virus infection, organ transplantation, or the administration of immunosuppressive drugs, was significantly more frequent in PCR-positive, RIBA3.0-indeterminate patients than in PCR-negative, RIBA3.0 indeterminate patients (P < 0.001) and PCR-positive patients with a positive RIBA3.0 result (P < 0.01). The distribution of HCV genotypes did not differ significantly between HCV RNA-positive patients with indeterminate or positive RIBA3.0 results. In conclusion, the prevalence of indeterminate RIBA3.0 patterns in virology laboratories is about 10%; in about half of these patients HCV replication is detected by PCR; the main factor responsible for indeterminate RIBA3.0 patterns could be immunosuppression, whereas HCV genotypes do not seem to play major role.
PMCID: PMC228735  PMID: 8748278
9.  Automated RIBA Hepatitis C Virus (HCV) Strip Immunoblot Assay for Reproducible HCV Diagnosis 
Journal of Clinical Microbiology  1998;36(2):387-390.
A comparison between the CHIRON RIBA hepatitis C virus (HCV) processor and manual systems was performed by using 88 specimens repeatedly reactive by the second-generation HCV enzyme-linked immunosorbent assay (ELISA) (HCV 2.0 ELISA) and 111 random specimens from volunteer donors. For the second-generation RIBA HCV strip immunoblot assay (SIA) (RIBA HCV 2.0 SIA), test results correlated strongly between the manual and the automated runs (kappa value, 0.937). For the RIBA HCV 3.0 SIA, the correlation of the test results was also high (kappa value, 0.899). Among the specimens with positive results by RIBA HCV 2.0 and 3.0 SIAs, there was a very strong concordance of the test results between the manual and the automated runs with regard to the reactive bands. Nine samples had discordant results between the manual and the automated runs; this was probably attributable to increased variability in antigen scores close to the cutoff values for both tests. Run-to-run and within-run testing by the CHIRON RIBA HCV Processor System showed a very low rate of conflicting values. In conclusion, the CHIRON RIBA HCV Processor System is capable of performing RIBA HCV 2.0 and 3.0 SIAs accurately with minimal operator involvement. In addition, the CHIRON RIBA HCV Processor System shows excellent reproducibility, with the potential for operator-to-operator and site-to-site variability being greatly reduced. Our data indicate that this novel methodology may be very useful for supplemental anti-HCV testing of specimens repeatedly reactive by ELISA in routine clinical assessments and epidemiologic evaluations.
PMCID: PMC104547  PMID: 9466746
10.  A pilot study on screening blood donors with individual-donation nucleic acid testing in China 
Blood Transfusion  2014;12(2):172-179.
Background
Nucleic acid amplification testing (NAT) is not yet obligatory in China for blood donor screening and the risk of enzyme immunoassay (EIA)-negative, NAT-reactive donations in Chinese blood donors has rarely been reported. The aim of this study was to screen a population of Chinese blood donors using a triplex individual-donation (ID)-NAT assay and assess the safety benefits of implementing NAT.
Materials and methods
Between 1st August, 2010 and 31st December, 2011 all donations at a Chinese blood centre were screened individually using the Procleix® Ultrio® assay, a multiplex NAT assay for the detection of hepatitis B virus (HBV) DNA, hepatitis C virus (HCV) RNA and human immunodeficiency virus-1 (HIV-1) RNA. All donations were also screened for HBsAg, anti-HIV and anti-HCV using two different EIA for each marker. Samples with discordant results between NAT and EIA were further tested with an alternative NAT assay (Cobas® TaqMan®). Potential yield cases (serologically negative/NAT-reactive donors) were further evaluated when possible.
Results
During the study period a total of 178,447 donations were screened by NAT and EIA, among which 169 HBV NAT yield cases (0.095%) were detected. No N AT yield cases were found for HIV-1 or HCV. For the HBV NAT yield cases, follow-up results showed that 11 (6.51%) were probable or confirmed HBV window period infections, 5 (2.96%) were chronic HBV carriers and 153 (90.53%) were probable or confirmed occult HBV infections. There was a statistically significant difference between the NAT-positive rates for first-time vs repeat donations (0.472% vs 0.146%, respectively; P<0.001).
Discussion
Our data demonstrate that the potential HBV yield rate was 1:1,056 for blood donations in the Zhejiang province of China. Implementation of NAT will provide a significant increment in safety relative to serological screening alone.
doi:10.2450/2013.0095-13
PMCID: PMC4039698  PMID: 24333061
nucleic acid amplification test; enzyme immunoassay; blood screening
11.  Transfusion transmittable infections – Seroprevalence among blood donors in a tertiary care hospital of Delhi 
Context:
Transfusion transmittable infections (TTI) continue to be a major threat to safe transfusion practices. Blood is one of the major sources of transmission of infectious diseases viz. human immunodeficiency virus (HIV), hepatitis B virus (HBV), hepatitis C virus (HCV), syphilis, malaria, and many other infections in India. Screening assays for the infectious diseases with excellent sensitivity and specificity helps to enhance the safety of the blood transfusions reducing the diagnostic window period as much as possible.
Aims:
The present study was designed to determine the seroprevalence of TTIs viz., HIV, HCV, and HBV, among the blood donors in Max Super Specialty Hospital, New Delhi, India based on dual testing strategy using high sensitive screening assays such as enhanced chemiluminescence assay and nucleic acid testing (NAT).
Materials and Methods:
A total of 41207 blood units collected from the donors (both voluntary and replacement donors) were screened for the TTI s, viz., anti HIV 1 and 2 antibody, anti HCV antibody, anti HBcore antibody, and HBsAg by enhanced chemiluminescence assay on VITROS® ECiQ immunodiagnostics system. NAT was performed using Roche Cobas® TaqScreen MPX assay, which can detect simultaneously HIV 1 (groups M and O), HIV-2, HCV, and HBV on Roche Cobas® s201 system.
Results:
The seroprevalence of HIV, HBsAg, anti HBcore antibody, and HCV based on enhanced chemiluminescence assay was found to be 0.25, 0.2, 7.06, and 0.7%, respectively. A total number of 6587 samples from July 2010 to December 2010 were tested on NAT, of which 3 samples were reactive for HBV in NAT; this was missed by enhanced chemiluminescence assay.
Conclusions:
Based on the seroprevalence study of infectious diseases viz., HIV, HBV, and HCV, we conclude that screening of blood and blood components by dual testing strategy using high sensitivity serological assay like enhanced chemiluminescence technology and NAT helps in detecting the potentially infectious blood units in all phases of infection, which aids in enhancing the safety of blood transfusion and reducing the potential risk of post-transfusion infection.
doi:10.4103/0973-6247.115566
PMCID: PMC3757770  PMID: 24014940
Seroprevalence; seroprevalence in blood donors; transfusion transmittable infections
12.  Indeterminate results of the second-generation hepatitis C virus (HCV) recombinant immunoblot assay: significance of high-level c22-3 reactivity and influence of HCV genotypes. 
Journal of Clinical Microbiology  1997;35(1):311-312.
A second-generation recombinant immunoblot assay (RIBA 2.0) is used in the United States to confirm infection with hepatitis C virus (HCV) in samples that are anti-HCV (enzyme immunoassay) positive. In some cases, indeterminate results of RIBA 2.0, which are defined as reactivity to a single antigen species or reactivity limited to two proteins derived from the same coding region of the HCV genome, are encountered. This study was performed to establish the significance of indeterminate RIBA 2.0 results in relation to HCV RNA detection, high positivity for the c22-3 band, and the HCV genotype as determined by direct DNA sequencing. Ninety-six samples with indeterminate RIBA 2.0 results were studied. HCV RNA was detected in 21 of 34 (62%) samples with high reactivity to c22-3 and in 8 of 62 (13%) samples with low reactivity to c22-3. The HCV genotype distribution in samples that were RIBA 2.0 indeterminate and HCV RNA positive was significantly different from that in samples of a control group with positive results for both the RIBA 2.0 and HCV PCR. These results suggest that highly positive c22-3 samples are likely to be associated with HCV viremia and that infection with less common HCV genotypes is more commonly associated with indeterminate RIBA 2.0 results.
PMCID: PMC229567  PMID: 8968936
13.  Prevalence and trends of markers of hepatitis B virus, hepatitis C virus and human Immunodeficiency virus in Argentine blood donors 
BMC Infectious Diseases  2014;14:218.
Background
Transfusion-transmitted infections are a major problem associated with blood transfusion. The aim of this study was to determine prevalence and trends of HBV, HCV and HIV in blood donors in Argentina.
Methods
A retrospective study was carried out in blood donors of 27 transfusion centers covering the whole country over a period of eight years (2004-2011). Serologic screening assays for HBsAg, anti-HBc, anti-HCV, and anti-HIV were performed in all centers and nucleic acid amplification testing (NAT) was performed in 2 out of the 27 centers.
Results
The 2,595,852 samples tested nationwide from 2004 to 2011 showed that the prevalence of HBsAg decreased from 0.336% to 0.198% (p < 0.0001), that of anti-HBc from 2.391% to 2.007% (p < 0.0001), that of anti-HCV from 0.721% to 0.460%, (p < 0.0001) and that of anti-HIV from 0.208% to 0.200 (p = 0.075). The prevalence of HBV, HCV and HIV was unevenly distributed among the different regions of the country. Two out of 74,838 screening- negative samples were positive in NAT assays (1 HIV-RNA and 1 HCV-RNA); moreover, HBV-DNA, HCV-RNA and HIV-RNA were detected in 60.29, 24.54 and 66.67% of screening-positive samples of the corresponding assays. As regards donors age, positive HBV-DNA and HCV-RNA donors were significantly older than healthy donors (46.6, 50.5 and 39.5 y respectively, p < 0.001).
Conclusions
Argentina has a low prevalence of HBsAg, anti-HCV and anti-HIV in blood donors, with a decreasing trend for HBsAg, anti-HBc and anti-HCV but not for anti-HIV over the last 8 years. The uneven distribution of transfusion-transmitted infections prevalence among the different regions of the country highlights the need to implement regional awareness campaigns and prevention. The discrepancy between samples testing positive for screening assays and negative for NAT assays highlights the problem of blood donors who test repeatedly reactive in screening assays but are not confirmed as positive upon further testing. The uneven distribution of age between healthy donors and NAT-positive donors could be related to changes in risks of these pathogens in the general population and might be attributed to a longer exposure to transmission risk factors in elderly people.
doi:10.1186/1471-2334-14-218
PMCID: PMC4018657  PMID: 24755089
Prevalence; Trend; Blood donors; HIV; HBV; HCV
14.  Need for Nucleic Acid Testing in Countries with High Prevalence of Transfusion-Transmitted Infections 
ISRN Hematology  2012;2012:718671.
Introduction. In India, family/replacement donors still provide more than 45% of the collected blood. With increasing voluntary blood donation and the still-prevalent infectious diseases in donors, we need to augment transfusion-transmitted infections (TTIs) testing before use. Our study was aimed to know the seroprevalence of TTIs among the donors of Rajasthan and the need for newer technologies like nucleic acid testing (NAT). Materials and Methods. Enhanced chemiluminescence immunoassay (ECi) was used for detection of HBsAg, anti-HIV, and anti-HCV in donor serum. 50% of the blood units which were negative on ECi were randomly selected and subjected to NAT testing for HBV, HCV, and HIV. Results. The total seroprevalence of TTIs is 2.62%. Of the randomly selected donor units negative by ECi, 8 turned out to be reactive on NAT testing: 4 were voluntary and 4 were family/replacement donors. Combined NAT yield (NAT reactive/seronegative) for HIV, HCV, and HBV was 0.034% (1 in 2972 donations). All the 8 reactive samples were positive for HBV DNA. Conclusion. In countries with a high prevalence of TTIs like India there are likely to be a significant number of window period donations that can be identified by NAT which may be implemented in blood centers allover India with serological testing to provide safe blood and cost alone should not be a deterrent to the government and implementing agencies.
doi:10.5402/2012/718671
PMCID: PMC3447329  PMID: 23008779
15.  A Novel Diagnostic Target in the Hepatitis C Virus Genome 
PLoS Medicine  2009;6(2):e1000031.
Background
Detection and quantification of hepatitis C virus (HCV) RNA is integral to diagnostic and therapeutic regimens. All molecular assays target the viral 5′-noncoding region (5′-NCR), and all show genotype-dependent variation of sensitivities and viral load results. Non-western HCV genotypes have been under-represented in evaluation studies. An alternative diagnostic target region within the HCV genome could facilitate a new generation of assays.
Methods and Findings
In this study we determined by de novo sequencing that the 3′-X-tail element, characterized significantly later than the rest of the genome, is highly conserved across genotypes. To prove its clinical utility as a molecular diagnostic target, a prototype qualitative and quantitative test was developed and evaluated multicentrically on a large and complete panel of 725 clinical plasma samples, covering HCV genotypes 1–6, from four continents (Germany, UK, Brazil, South Africa, Singapore). To our knowledge, this is the most diversified and comprehensive panel of clinical and genotype specimens used in HCV nucleic acid testing (NAT) validation to date. The lower limit of detection (LOD) was 18.4 IU/ml (95% confidence interval, 15.3–24.1 IU/ml), suggesting applicability in donor blood screening. The upper LOD exceeded 10−9 IU/ml, facilitating viral load monitoring within a wide dynamic range. In 598 genotyped samples, quantified by Bayer VERSANT 3.0 branched DNA (bDNA), X-tail-based viral loads were highly concordant with bDNA for all genotypes. Correlation coefficients between bDNA and X-tail NAT, for genotypes 1–6, were: 0.92, 0.85, 0.95, 0.91, 0.95, and 0.96, respectively; X-tail-based viral loads deviated by more than 0.5 log10 from 5′-NCR-based viral loads in only 12% of samples (maximum deviation, 0.85 log10). The successful introduction of X-tail NAT in a Brazilian laboratory confirmed the practical stability and robustness of the X-tail-based protocol. The assay was implemented at low reaction costs (US$8.70 per sample), short turnover times (2.5 h for up to 96 samples), and without technical difficulties.
Conclusion
This study indicates a way to fundamentally improve HCV viral load monitoring and infection screening. Our prototype assay can serve as a template for a new generation of viral load assays. Additionally, to our knowledge this study provides the first open protocol to permit industry-grade HCV detection and quantification in resource-limited settings.
Christian Drosten and colleagues develop, validate, and make openly available a prototype hepatitis C virus assay based on the conserved 3' X-tail element, with potential for clinical use in developing countries.
Editors' Summary
Background.
About 3% of the world's population (170 million people) harbor long-term (chronic) infections with the hepatitis C virus (HCV) and about 3–4 million people are newly infected with this virus every year. HCV—a leading cause of chronic hepatitis (inflammation of the liver)—is spread through contact with the blood of an infected person. Globally, the main routes of transmission are the use of unscreened blood for transfusions and the reuse of inadequately sterilized medical instruments, including needles. In affluent countries, where donated blood is routinely screened for the presence of HCV, most transmission is through needle sharing among drug users. The risk of sexual and mother-to-child transmission of HCV is low. Although HCV infection occasionally causes an acute (short-lived) illness characterized by tiredness and jaundice (yellow eyes and skin), most newly infected people progress to a symptom-free, chronic infection that can eventually cause liver cirrhosis (scarring) and liver cancer. HCV infections can be treated with a combination of two drugs called interferon and ribavirin, but these drugs are expensive and are ineffective in many patients.
Why Was This Study Done?
An effective way to limit the global spread of HCV might be to introduce routine screening of the blood that is used for transfusions in developing countries. In developed countries, HCV screening of blood donors use expensive, commercial “RT-PCR” assays to detect small amounts of HCV ribonucleic acid (RNA; HCV stores the information it needs to replicate itself—its genome—as a sequence of “ribonucleotides”). All the current HCV assays, which can also quantify the amount of viral RNA in the blood (the viral load) during treatment, detect a target sequence in the viral genome called the 5′-noncoding region (5′-NCR). However, there are several different HCV “genotypes” (strains). These genotypes vary in their geographical distribution and, even though the 5′-NCR sequence is very similar (highly conserved) in the common genotypes (HCV genotypes 1–6), the existing assays do not detect all the variants equally well. This shortcoming, together with their high cost, means that 5′-NCR RT-PCR assays are not ideal for use in many developing countries. In this study, the researchers identify an alternative diagnostic target sequence in the HCV genome—the 3′-X-tail element—and ask whether this sequence can be used to develop a new generation of tests for HCV infection that might be more appropriate for use in developing countries.
What Did the Researchers Do and Find?
The researchers determined the RNA sequence of the 3′-X-tail element in reference samples of the major HCV genotypes and showed that this region of the HCV genome is as highly conserved as the 5′-NCR. They then developed a prototype X-tail RT-PCR assay and tested its ability to detect small amounts of HCV and to measure viral load in genotype reference samples and in a large panel of HCV-infected blood samples collected in Germany, the UK, Brazil, South Africa, and Singapore. The new assay detected low levels of HCV RNA in all of the genotype reference samples and was also able to quantify high RNA concentrations. The viral load estimates it provided for the clinical samples agreed well with those obtained using a commercial assay irrespective of the sample's HCV genotype. Finally, the X-tail RT-PCR assay gave similar results to a standard assay at a fraction of the cost when used to measure viral loads in a Brazilian laboratory in an independent group of 127 patient samples collected in Brazil.
What Do These Findings Mean?
These findings suggest that the HCV 3′-X-tail element could provide an alternative target for screening blood samples for HCV infection and for monitoring viral loads during treatment, irrespective of HCV genotype. In addition, they suggest that X-tail RT-PCR assays may be stable and robust enough for use in laboratories in emerging countries. Overall, these findings should stimulate the development of a new generation of clinical HCV assays that, because the protocol used in the X-tail assay is freely available, could improve blood safety in developing countries by providing a cheap and effective alternative to existing proprietary HCV assays.
Additional Information.
Please access these Web sites via the online version of this summary at http://dx.doi.org/10.1371/journal.pmed.1000031.
The World Health Organization has a fact sheet about hepatitis C (in English and French)
The US Centers for Disease Control and Prevention provides information on hepatitis C for the public and for health professionals (information is also available in Spanish)
The US National Institute of Diabetes and Digestive and Kidney Diseases provides basic information on hepatitis C (in English and Spanish)
The MedlinePlus Encyclopedia has a page on hepatitis C; MedlinePlus also provides links to further information on hepatitis C (in English and Spanish)
doi:10.1371/journal.pmed.1000031
PMCID: PMC2637920  PMID: 19209955
16.  Comparison of Second- and Third-Generation Enzyme Immunoassays for Detecting Antibodies to Hepatitis C Virus 
Journal of Clinical Microbiology  2002;40(5):1656-1659.
Supplemental assays, such as recombinant immunoblot assays (RIBA), are used to confirm detection of antibodies to hepatitis C virus (HCV). However, due to their expense, they are not widely used in developing countries. The purpose of our study was to compare the results of second- and third-generation (G2 and G3, respectively) enzyme immunoassays (EIAs) and to resolve discordant results by using a supplemental assay to assess the reliability of G2 and G3 EIAs to confirm anti-HCV antibody-positive results. We performed both G2 and G3 EIAs for anti-HCV antibodies on 1,134 serum samples collected during the 2nd year of a longitudinal community-based study in Egypt; 35 samples with discordant results were tested by Abbott Laboratories Micro-Particle Immunoassay (M-EIA) and RIBA. Viremia was determined with an in-house nested reverse transcriptase PCR (RT-PCR) to detect HCV RNA. Concordance between the two assays (G2/G3) was 96.9%; 87 (7.7%) samples were positive and 1,012 (89.2%) were negative by both assays. For 17 samples, the discordant results were G2 assay negative and G3 assay positive, and for 18 samples, the discordant results were G2 assay positive and G3 assay negative. Among the 17 G2 assay-negative and G3 assay-positive samples, 15 were M-EIA positive and 7 were PCR positive. Among the 18 G2 assay-positive and G3 assay-negative samples, 2 were M-EIA positive and none were PCR positive. RIBA results from 24 discordant samples showed 87.5% agreement with the G3 EIA, 12.5% agreement with the G2 EIA, and 95.8% agreement with M-EIA. Eleven samples were indeterminate by RIBA and excluded from this analysis. Based on RIBA results, the sensitivity of the G3 EIA was 99%, compared to 89.8% for the G2 EIA, while the specificity of the G3 EIA was 99.8%, compared to 98.9% for the G2 EIA. These results show that the reliability of the G3 EIA in screening these sera is excellent, and the G3 assay can be used in the absence of supplemental tests where resources are limited. RIBA appears not to have advantages over the less expensive M-EIA screening assay. The main disadvantage of RIBA is the occurrence of indeterminate results, especially among problematic samples. Samples giving discordant results in multiple assays are often indeterminate with the RIBA.
doi:10.1128/JCM.40.5.1656-1659.2002
PMCID: PMC130934  PMID: 11980937
17.  Study on reliability of commercially available hepatitis C virus antibody tests. 
Journal of Clinical Microbiology  1995;33(3):620-624.
The serodiagnosis of hepatitis C virus (HCV) infection was analyzed by a recombinant immunoblot assay (RIBA) with recombinant proteins encoded by the viral RNA isolated from our patients in Hamburg, Germany. The HCV RNA was amplified by PCR, and proteins encoded by the viral core and the NS3, NS4, and NS5 regions were expressed subsequently in Escherichia coli. The results obtained with our UKE RIBA were compared with the results of the Abbott HCV second-generation enzyme immunoassay (EIA). Serum samples from 270 patients, which were sent to us on the suspicion of HCV hepatitis and which were negative for hepatitis A virus and hepatitis B virus antibodies, were examined. In 227 cases (84.1%), there were identical positive (204 cases, 75.6%) or negative (23 cases, 8.5%) results in both tests. In 32 cases (11.9%), the reactive Abbott second-generation HCV EIA results could not be confirmed by the UKE RIBA and the HCV PCR. In follow-up studies conducted over 1 year, these results did not change. In three cases (1.1%), the UKE RIBA presented a positive result while the Abbott second-generation HCV EIA was negative. All three cases were positive in the HCV PCR and showed seroconversion in an HCV EIA 4 to 6 weeks later. In addition, 33 patient serum samples were examined by UKE RIBA in parallel with the Ortho RIBA 2.0. In three cases (9.1%), a positive Ortho RIBA 2.0 result could not be confirmed by the UKE RIBA and the HCV PCR. All three patients were free of complaints. The UKE RIBA showed also a smaller number of indeterminate results (3.0%) than the Ortho RIBA 2.0 (24.2%). This comparison study demonstrates that the commercially available HCV antibody tests should be further improved.
PMCID: PMC228001  PMID: 7751366
18.  Confirmation of second generation anti-hepatitis C virus enzyme immunoassays by antigenic cross-validation. 
Journal of Clinical Pathology  1992;45(10):917-920.
AIM: To determine if a scheme for validating enzyme immunoassay (EIA) results could be devised that did not require costly and methodically elaborate supplemental assays. METHODS: Samples (n = 525) from patients with haemophilia A, leukaemia, and chronic liver disease and at increased risk of hepatitis C virus infection were tested by EIA-1 (Ortho Diagnostics), an assay which uses recombinant HCV fusion proteins as antigens, and by EIA-2 (United Biomedical), an assay based on synthetic HCV oligopeptide antigens. RESULTS: Samples (n = 193) were repeatedly reactive in both EIAs. Of these, 190 (98%) yielded reactivities in both of two supplemental assays used, one an immunoblot assay (RIBA) using recombinant HCV polypeptides similar to EIA-1 antigens, and the other a neutralisation EIA (EIA-2N) based on antigenic competition with HCV peptides similar to EIA-2 antigens. The three samples not reactive in supplemental tests exhibited low EIA optical density (OD) values (signal/cutoff ratios of less than 3). Hence, all specimens reactive and yielding high OD values in both EIAs were also reactive in supplemental assays. Twenty four samples were reactive in EIA-1 only and nine (38%) of these were reactive in RIBA. Fourteen of the 15 (93%) specimens reactive in EIA-1 but not RIBA were derived from patients with chronic liver dysfunction. Two samples were reactive in EIA-2 only, of which one was reactive in EIA-2N and none in RIBA. CONCLUSIONS: Compared with EIA-2, EIA-1 yielded more validated reactive samples and resulted in more non-validated reactivities. It is therefore suggested that for clinical diagnosis: (i) EIA-1 be used for anti-HCV testing and EIA-2 for validation of EIA-1 reactivities; (ii) samples concordantly reactive in EIA-1 and EIA-2 and displaying high OD readings be considered HCV antibody positive without supplemental testing; (iii) supplemental testing by RIBA be limited to samples reactive in EIA-1 but equivocal or unreactive in EIA-2 and those concordantly reactive but exhibiting low absorbance readings.
PMCID: PMC495067  PMID: 1331199
19.  Anti-HBc screening in Indian blood donors: Still an unresolved issue 
AIM: To study the seroprevalence of antibody to hepatitis B core antigen (anti-HBc) in healthy blood donors negative for HBsAg and to evaluate whether anti-HBc detection could be adopted in India as a screening assay for HBV in addition to HBsAg.
METHODS: A total of 1700 serum samples collected from HBsAg-negative healthy blood donors were tested for the presence of anti-HBc antibody (IgM + IgG). All samples reactive for anti-HBc antibody were then investigated for presence of anti-HBs and for liver function tests (LFTs). One hundred serum samples reactive for anti-HBc were tested for HBV DNA by PCR method.
RESULTS: Out of 1700 samples tested, 142 (8.4%) blood samples were found to be reactive for anti-HBc. It was significantly lower in voluntary (6.9%) as compared to replacement donors (10.4%, P = 0.011). Seventy-two (50.7%) anti-HBc reactive samples were also reactive for anti-HBs with levels > 10 mIU/mL and 70 (49.3%) samples were non-reactive for anti-HBs, these units were labeled as anti-HBc-only. These 142 anti-HBc reactive units were also tested for liver function test. HBV DNA was detected in only 1 of 100 samples tested.
CONCLUSION: Keeping in view that 8%-18% of donor population in India is anti-HBc reactive, inclusion of anti-HBc testing will lead to high discard rate. Anti-HBs as proposed previously does not seem to predict clearance of the virus. Cost effectiveness of introducing universal anti-HBc screening and discarding large number of blood units versus considering ID NAT (Individual donor nuclic acid testing) needs to be assessed.
doi:10.3748/wjg.14.5327
PMCID: PMC2744065  PMID: 18785287
Hepatitis B core antigen; Hepatitis B surface antigen; Hepatitis B virus; Transfusion-associated hepatitis B virus; Blood donors
20.  Hepatitis C virus in sickle cell disease. 
PURPOSE: To determine the prevalence of hepatitis C virus antibodies (anti-HCV) in patients with sickle cell disease. PATIENTS AND METHODS: Between 1983 and 2001, 150 patients from the Howard University Hospital Center for Sickle Cell Disease were screened for HCV antibody (52% women, 48% men, mean age 34 years). Frozen serum samples from 56 adult sickle cell patients who had participated in previous surveys (1983-92) of HIV and HTLV-1 serology and who were tested in 1992 for anti-HCV antibody--when commercial ELISA test (Ortho) became available--were included in this paper. Of the 150 patients in the study, 132 had sickle cell anemia genotype (SS), 15 had sickle cell hemoglobin-C disease (SC) and three had sickle beta thalassemia. Clinical charts were reviewed for history of blood transfusion, IV drug abuse, homosexuality, tattooing, iron overload, and alcohol abuse. RESULTS: Antibodies to HCV were detected in 53 patients (35.3%). Of the 55 patients who had frozen serum samples tested in 1992, 32 (58%) were reactive for anti-HCV, while only 21 of the 95 patients (22%) tested after 1992 were positive for HCV antibodies (P<0.001). Thirty-nine of 77 patients (51%) who received more than 10 units of packed red blood cells were positive for HCV antibody, and only 14 of 61 patients (23%) who received less than 10 units of packed red blood cells transfusion were positive for HCV antibodies (P<0.001). None of the 12 patients who never received transfusion were positive for HCV antibody. In the 53 anti-HCV positive patients, the mean alanine amino-transferase (ALT) value was 98- and 81 U/L, respectively, for males and females. These values were normal for the HCV-antibody negative patients. The aspartate amino-transferase (AST) and the total bilirubin were also higher in the anti-HCV positive patients compared to patients in the anti-HCV negative group. Forty-four patients (57.1%) who were transfused more than 10 units developed iron overload defined by a serum ferritin level higher than 1,000 ng/ml. A total of 20 of the patients with iron overload underwent liver biopsies. Seven of these 20 patients (35%) were HCV positive. These patients often had more severe liver disease and higher degree of iron deposition. CONCLUSION: The prevalence of HCV antibody and iron overload is directly related to the number of blood transfusions in patients with sickle cell disease. The prevalence of HCV infection has decreased significantly, since blood donor screening for HCV became available. Chronic HCV infection and iron overload place sickle cell patients at risk for significant liver disease.
PMCID: PMC2594496  PMID: 14620705
21.  Prevalence of hepatitis C virus infection among human immunodeficiency virus-1-infected pregnant women in Malawi: The BAN study☆ 
Background
In Sub-Saharan Africa, prevalence estimates of hepatitis C virus (HCV) vary widely.
Objectives
To assess the prevalence of HCV infection among HIV-infected, pregnant women screened for a large clinical trial in Lilongwe, Malawi.
Study design
Plasma from 2041 HIV-infected, pregnant women was screened for anti-HCV IgG using a chemiluminiscent immunometric assay (CIA). Specimens with a signal-cut-off ratio ≥ 1.00 were considered reactive and those with S/Co ratio < 1.00 non-reactive. All CIA-reactive specimens were tested by a recombinant immunoblot assay (RIBA) for anti-HCV and by PCR for HCV RNA.
Results
Of 2041 specimens, 110 (5.3%, 95% CI: 4.5–6.5%) were CIA reactive. Of the 109 CIA reactive specimens available for RIBA testing, 2 (1.8%) were positive, 28 (25.7%) were indeterminate, and 79 (72.5%) were negative. All CIA-reactive specimens were HCV RNA negative (n = 110). The estimated HCV prevalence based on the screening assay alone was 5.3%; based on supplemental RIBA testing, the status of HCV infection remained indeterminate in 1.4% (28/2040, 95% CI: 0.1–2.0) and the prevalence of confirmed HCV infections was 0.1% (2/2040, 95% CI: 0–0.4%).
Conclusions
HCV seroprevalence among HIV-infected, pregnant women in Malawi confirmed by supplemental RIBA HCV 3.0 is low (0.1%); CIA showed a high false-reactivity rate in this population.
doi:10.1016/j.jcv.2012.05.003
PMCID: PMC3652577  PMID: 22658797
HIV; HCV; Pregnant women; Malawi
22.  Antibody to hepatitis C virus in risk groups in Canada 
The prevalence of antibodies against hepatitis C virus (HCV) was studied in hemophiliacs, hemodialysis patients, intravenous drug abusers, female prisoners, homosexuals, individuals with no markers of recent hepatitis A or B virus infections and normal individuals (federal public servants), by an enzyme immunoassay (Ortho Diagnostic Systems Inc). Repeat positive samples were further tested by recombinant immunoblot assay (riba) HCV (Chiron Corp, California). The number of samples positive for antibodies to HCV (anti-HCV) was higher with enzyme immunoassay than by riba HCV in most cases. A high prevalence of anti-HCV was detected in hemophiliacs by both enzyme immunoassay (68.8%) and riba HCV (53.7%). Among intravenous drug abusers and female prisoners the prevalence rates for anti-HCV were 42.8% and 29.8%, respectively, by riba HCV; the results with enzyme immunoassay were only slightly higher. The prevalence rate was also high by both tests (54.2%) in hemodialysis patients’ sera taken during 1980–82, when many cases of non-A,non-B hepatitis were suspected in this group. In contrast, only 14.1% of sera taken during 1990 were positive by riba HCV. In individuals with no markers of recent hepatitis A or B infections, 13.4% were positive by enzyme immunoassay, whereas only 4.5% were reactive by riba HCV. The lowest prevalence was seen in homosexuals (2.3%) and normal individuals (1.2%) by riba HCV. These results indicate a high prevalence of anti-HCV in high risk groups tested in Canada.
PMCID: PMC3307416  PMID: 22451759
Enzyme immunoassay; Hepatitis C virus; Non-A,non-B hepatitis
23.  Clinical significance of serum hepatitis C virus (HCV) RNA as marker of HCV infection. 
Journal of Clinical Microbiology  1994;32(12):3008-3012.
We have evaluated the clinical significance of hepatitis C virus (HCV) RNA determination by analyzing a group of 221 hospitalized patients with abnormal liver function tests. Serum HCV RNA was detected by "nested" PCR amplification followed by nonisotopic hybridization. Of the 200 (90.5%) patients with anti-HCV-positive enzyme-linked immunosorbent assay results, 152 (76%) were RIBA reactive, 47 (23.5%) had indeterminate results, and 1 (0.5%) was nonreactive. Of the 180 (90%) patients positive for anti-HCV and HCV RNA, 138 (76.7%) were RIBA reactive and 42 (23.3%) were RIBA indeterminate. The pattern of RIBA reactivity did not correlate with the presence of HCV RNA. Elevated alanine aminotransferase levels were associated neither with the presence of viremia nor with the RIBA pattern. Histological findings consistent with non-A non-B hepatitis correlated with the presence of HCV RNA but not with the RIBA pattern. HCV RNA was detected in 11 of 21 (52.4%) anti-HCV-negative patients. These 11 patients were either immunosuppressed or in the prodromic phase of acute hepatitis C. Circulating HCV RNA can therefore be described as being predictive of virus-induced liver damage in anti-HCV-positive patients and may be useful in the diagnosis of HCV infection in anti-HCV-negative immunosuppressed patients or in those with early acute infection.
Images
PMCID: PMC264216  PMID: 7883891
24.  Analysis of sera indeterminate by Ortho-HCV RIBA-2 by using three confirmatory assays for anti-hepatitis C virus antibody. 
Journal of Clinical Microbiology  1994;32(9):2071-2075.
The diagnostic performances of three commercially available recombinant immunoblot assays (RIBAs) for anti-hepatitis C virus antibody were evaluated on 50 ORTHO-HCV RIBA-2 (RIBA-2)-indeterminate serum samples. Concordant interpretations were obtained with the three tests in 60% of the samples, with 56% positive, 2% indeterminate, and 2% negative results. Considering test performance in regard to the number of remaining indeterminate results, analyzing sera by RIBA-3, INNO-LIA HCV Ab III, and DECISCAN HCV reduced the number of samples reacting indeterminately to 40, 6, and 8%, respectively. The three serum samples classified as indeterminate in the INNO-LIA HCV Ab III as well as three of four serum samples interpreted as indeterminate in the DECISCAN HCV and 16 of 20 samples classified as indeterminate in the RIBA-3 were hepatitis C virus RNA positive by PCR. This study clearly shows the good performance of the three tests as confirmatory assays compared with that of the RIBA-2. However, according to the manufacturers' criteria of positivity, the INNO-LIA HCV Ab III and DECISCAN HCV appeared to be more suitable than the RIBA-3 for interpreting serum samples found indeterminate in the RIBA-2.
PMCID: PMC263944  PMID: 7529247
25.  Diverse patterns of recognition of hepatitis C virus core and nonstructural antigens by antibodies present in Egyptian cancer patients and blood donors. 
Journal of Clinical Microbiology  1996;34(11):2665-2669.
Serum samples from 429 cancer patients, 82 unpaid blood donors, and 74 paid blood donors were tested for hepatitis C virus (HCV) markers in two commercially available enzyme immunoassays (EIAs). A total of 229 of 429 (53.4%) cancer patients were positive by the two EIAs. A total of 34 of 156 (21.8%) of the blood donors were positive by the EIAs, with a higher prevalence among paid blood donors (20/74; 27%) compared with that among the unpaid blood donors (14 of 82; 17%). EIA-positive sera were tested for confirmation of the results in an immunoblot assay (LiaTek) in which reactivities to four synthetic peptides representing the HCV core protein and two synthetic peptides representing nonstructural proteins 4 and 5 were measured. Of 243 first and/or second EIA-positive samples from cancer patients, 188 (77.2%) were confirmed to be positive in the synthetic peptide immunoblot. A total of 33 of 35 (94.3%) blood donor samples were confirmed to be positive. A great diversity in reactivity patterns was seen. However, all sera from the group of paid blood donors were exclusively reactive to core peptides 1 and 2. A subset of LiaTek assay-positive samples were tested by the four-antigen RIBA-2 assay. The sera from the paid blood donors were all nonreactive. A subset of the LiaTek-positive sera was analyzed for the presence of the HCV genome by reverse transcriptase-PCR. Eleven of the 20 serum samples with reactivity to LiaTek core peptides 1 and 2 only were HCV reverse transcriptase-PCR positive, as were the majority of the sera with other reactivity patterns by the LiaTek assay. The results confirm the very high prevalence of HCV infection in Egypt. Furthermore, the results indicate that there is circulating in Egypt, particularly in the group of blood donors paid for their donation, an HCV variant which elicits an immune response that is not detected by the RIBA-2 assay.
PMCID: PMC229382  PMID: 8897161

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