Use of seroconversion panels to estimate delay in detection of anti-human immunodeficiency virus antibodies by enzyme-linked immunosorbent assay of pooled compared to singleton serum samples

The transfusion of unsafe blood worldwide accounts for 5 to 15% of new human immunodeficiency virus (HIV) infections, most of which occur in sub-Saharan Africa. While developed countries now apply PCR testing of pooled samples, some developing countries still do not have universal screening policies. More efficient low-cost procedures for the screening of pooled samples have the potential to encourage mass screening efforts in resource-poor settings. The aim of this study was to estimate the delay in the detection of HIV antibodies in pooled serum samples compared to that in singleton serum samples by enzyme-linked immunosorbent assay (ELISA) and to evaluate the risk of transfusion-transmitted HIV infection during the window period. Serial blood samples obtained from five HIV seroconversion panels were mixed with HIV-seronegative blood samples to create pools of 6, 12, 16, 24, 32, and 48 samples. The delay in detection of the first anti-HIV antibody-positive sample in tests with pooled samples was calculated for each pool size and compared to that obtained by testing of singleton samples and statistically evaluated by a robust log-linear regression analysis. The risk of a false-negative (FN) result caused by dilution was estimated by use of the incidence risk/window period model. The additional risk of transmission related to ELISA screening of pooled samples for HIV did not exceed 9% of the current risk of an FN result (estimated to be 1/1,067,000). The countries with virus prevalence rates in donors of less than 15% are expected to save up to 30% in the number of tests. ELISA screening of pooled samples could be considered in settings where the testing of blood supplies for HIV is not routinely done.     

The accuracy of an alternative confirmatory strategy for detection of antibodies to HIV-1: experience from a regional laboratory in Kagera, Tanzania.

BACKGROUND: Constant improvement of HIV tests often results in withdrawal of poorer quality tests by the manufacturing companies. It is thus often necessary to evaluate new HIV testing kits and modify the existing testing strategies. OBJECTIVES: To evaluate an alternative HIV antibody testing strategy which involves consecutive testing of sera by two enzyme-linked immunosorbent assays (ELISA), which both are recombinant antigen-based but utilise different test principles, followed by re-testing of sera giving discordant results. STUDY DESIGN: Sera (n = 1558) from a cross-sectional study of the HIV-1 seroprevalence in the Kagera region of Tanzania were tested using two ELISAs in parallel: Enzygnost anti-HIV-1/2 plus and Wellcozyme HIV-1 recombinant. Western blot analysis was done on all concordantly reactive and repeatedly discordant reactive samples as well as on 10% of concordantly ELISA negative sera. RESULTS: Two hundred and four sera (13.1%) were confirmed HIV-1-antibody positive. Both ELISAs had a sensitivity of 100%. The specificities of the ELISAs at initial and repeated testing were 99.8 and 99.9%, respectively, for Enzygnost and 97.7 and 99.5%, respectively, for Wellcozyme. None of the sera was concordantly false positive in both ELISAs. The mean ratio of the optical density of a sample to the cut off value of the test run (OD/CO ratio) was lower for samples giving false positive reactions than for confirmed HIV-1-antibody-positive samples. It is therefore important to interpret with caution HIV antibody ELISA test results on samples giving low OD/CO ratios. None of 10% of randomly selected concordantly ELISA negative sera gave a positive Western blot reaction. CONCLUSIONS: This field evaluation of an HIV antibody testing strategy involving the use of a recombinant antigen-based sandwich ELISA (Enzygnost) followed by a recombinant antigen-based competitive ELISA (Wellcozyme) showed that it had a sensitivity and specificity of 100%.     

Combining pooling and alternative algorithms in seroprevalence studies.

Data from two seroprevalence studies and one comparative study of confirmatory algorithms were used to compare the costs and sensitivities of six algorithms for determining seropositivity to human immunodeficiency virus (HIV). We evaluated confirmatory strategies by using the CBC Recombigen HIV enzyme immunoassay (EIA; Cambridge BioScience, Worcester, Mass.) and immunoblotting followed by radioimmunoprecipitation assay to confirm indeterminate immunoblotting results with and without pooling of samples during screening. The least expensive algorithm was that in which sera were pooled during screening and EIA was used to confirm positive test results. The cost savings associated with this confirmatory test were greater when the prevalence of HIV infection was higher. Savings from pooling of sera for screen testing diminished as HIV prevalence increased. The sensitivity and specificity of EIA with respect to immunoblotting and radioimmunoprecipitation assay were estimated to be 0.9992 and 0.9977, respectively. We found that the implementation of pooling during screening and the use of EIA as the confirmatory test do not affect the statistical reliability of estimates of seropositivity but do result in considerable cost savings.     

Pooling of sera for human immunodeficiency virus (HIV) testing: an economical method for use in developing countries.

To determine whether donated blood samples in African countries could be pooled, then tested for the presence of human immunodeficiency virus (HIV) antibodies with a single test without loss of accuracy, a single test on five pooled samples was used, followed by individual testing of positive pools. This resulted in no loss of either sensitivity or specificity. Pooling 10 samples resulted in a loss of sensitivity for low antibody titre specimens. Pooling reduced the costs of screening by 70% and time needed for analysis. It is concluded that pooling of five samples for HIV screening may result in a substantial reduction in costs; in countries where the prevalence of HIV is higher than the 2-3% found in Zimbabwean donors, however, savings may not be as great.     

Evaluating the efficiency of specimen (sample) pooling for real-time PCR based diagnosis of COVID-19

PurposeThis study is aims at evaluating the efficacy and sensitivity of specimen pooling for testing of SARS-CoV-2 virus to determine the accuracy, resource savings, and identification of borderline positive cases without impacting the accuracy of the testing.MethodThis study was conducted between August and October 2020, we performed COVID-19 testing by RT-PCR on the samples from varying prevalence of rural population (non-hot spot) referred to COVID laboratory, in the first step, the samples were collated into pools of 5 or 10. These pools were tested by RT-PCR. Negative pools were reported as negative whereas positive pools of 5 and 10 were then de-convoluted and each sample was tested individually.ResultsIn the present study, we tested 1580 samples in 158 pools of 10 and 17,515 samples in 3503 pools of 5. Among 10 samples pool, 11 (13%) pools flagged positive in the first step. In the second step, among 11 pools (110 samples) de-convoluted strategy was followed in which 10 individual samples came positive. Among 5 samples pool, 164 (13%) pools flagged positive in the first step. In the second step, among 164 pools (820 samples) de-convoluted strategy was followed in which 171 individual samples came positive. The pooled sample testing strategy saves substantial resources and time during surge testing and enhanced pandemic surveillance. This approach requires around 76%–93% fewer tests in low to moderate prevalence settings and group sizes up to 5–10 in a population, compared to individual testing.ConclusionPooled sample RT- PCR analysis strategies can save substantial resources and time for COVID-19 mass testing in comparison with individual testing without compromising the quality of outcome of the test. In particular, the pooled sample approach can facilitate mass screening in the early asymptomatic stages of COVID-19 infections.     

Diagnostic testing for HIV type 1 RNA in seronegative blood

We studied the feasibility of routine diagnostic testing for HIV-1 RNA at a publicly funded testing site. HIV-1 RNA was determined with a commercial polymerase chain reaction assay in pooled seronegative blood samples submitted for HIV testing to a public health laboratory. Recovery of HIV-1 RNA from the samples was estimated as at least 8% of viral RNA that was found in freshly prepared plasma. We estimated that screening for HIV-1 RNA in serum pools would result in the identification of blood specimens from more than 95% of acutely infected patients. The frequency of HIV-1 RNA in seronegative blood samples was estimated to be between 19 and 601 per 10(6) submitted specimens. The ratio of HIV-1 RNA positive and seronegative samples to specimens with HIV-1 antibodies confirmed by Western blot was estimated to be between 0.2% and 6.6%. The reagent costs for identifying 1 HIV-infected blood sample were 10-fold higher with the commercially available HIV-1 RNA assay compared with the HIV antibody enzyme-linked immunosorbent assay. Diagnostic testing for HIV-1 RNA may be warranted in high-risk populations since acutely infected patients may benefit most from anti-retroviral therapy and are thought to contribute disproportionately to the HIV epidemic.     

Sensitivity and specificity of pooled versus individual sera in a human immunodeficiency virus antibody prevalence study.

We evaluated the efficacy of testing pooled versus individual sera for the detection of human immunodeficiency virus antibody. A total of 5,000 individual specimens and 500 pools of 10 specimens each were assayed by an enzyme-linked immunosorbent assay. There was complete agreement in human immunodeficiency virus enzyme-linked immunosorbent assay reactivity for pooled versus individual specimens. An estimated savings of 60 to 80% (labor and supplies) can be realized dependent upon pooling and assay format.     

Routine antenatal screening for hepatitis B using pooled sera: validation and review of 10 years experience.

AIMS: To validate the sensitivity of universal antenatal screening for hepatitis B surface antigen (HBsAg) by testing pools of 10 sera, and to review 10 years' experience using this method. METHODS: 66,945 antenatal patients were tested between 1986 and 1996 using the pooled method. All sera from 1996 (n = 6050) were retrieved and retrospectively tested individually. An in vitro determination of the effect of pooling on sensitivity was performed by checkerboard neutralisation assay. RESULTS: 26 HBsAg positive women were detected by universal screening over 10 years; 12 had non-European surnames and five had known risk factors for hepatitis B infection. High titre anti-HBs sera in the pool reduced the sensitivity of the HBsAg assay, though the effect was only significant at low levels of HBsAg carriage. CONCLUSIONS: The prevalence of hepatitis B is extremely low in the antenatal population served by Plymouth PHL. Pooling is unlikely to reduce sensitivity enough to lead to significant preventable vertical transmission, and is a cost-effective and valid strategy in areas of low seroprevalence.     

Evaluation of a rapid test for detection of antibodies against human immunodeficiency virus (HIV)

A new test for rapid detection of HIV antibodies (HIVCHEK) was evaluated. A total of 107 sera were examined. Of these, 60 were from healthy blood donors and the rest from a serum panel containing HIV positives and samples with false positive reactions in ELISA and unspecific bands in the western blot (WB). There was close agreement (100%) between the methods. The specificity of the HIVCHEK was 100% in this study. The sensitivity was acceptably high based on testing of consecutive serum samples from two seroconversion patients. The test was rapid and easy to perform, and no extra equipment was needed.     

Evaluation of the Recombigen HIV-1 Latex Agglutination Test.

The Recombigen HIV-1 Latex Agglutination (LA) Test was recently licensed by the U.S. Food and Drug Administration for use as a rapid screening assay for human immunodeficiency virus type 1 (HIV-1) antibodies. However, its performance in various settings and in different populations has not been firmly established. Consequently, we evaluated the test in the Cleveland Clinic Retrovirus Laboratory, a regional reference laboratory for HIV diagnostic testing and a testing laboratory for the Ohio Department of Health Anonymous HIV Testing and Counseling Program. Serum samples from 93 individuals presumed to be at high risk for HIV infection were evaluated. The sera were initially tested for HIV antibodies by enzyme-linked immunosorbent assay (ELISA). All repeatedly reactive sera were subjected to confirmatory Western blot (WB; immunoblot) testing. Of 97 serum specimens tested (5 were from one seroconverter), 44 were repeatedly reactive by ELISA and 53 were nonreactive. Of the reactive serum specimens, 31 were confirmed positive and 12 were indeterminate by WB. All of the sera were coded and then retested by the LA test. Of 53 serum specimens nonreactive by ELISA, 51 were also nonreactive in the LA test. Of the 44 serum specimens reactive by ELISA, 16 were nonreactive by LA; however, 3 of the latter were WB positive. No serum specimen with an ELISA ratio (specimen optical density/cutoff optical density) of less than 2.1 scored reactive in the LA test. The LA test was positive for only two of five consecutive serum specimens from a seroconverter despite the fact that all but the earliest of these were ELISA reactive and WB positive. Although the LA test appears to be an adequate first-line screening test when appropriately used according to the directions of the manufacturer, our data suggest that occasional sera with low levels of reactivity by ELISA may not be readily detected as reactive by the LA test.     

Seroprevalence of severe acute respiratory syndrome coronavirus 2 in Slovenia: results of two rounds of a nationwide population study on a probability-based sample,challenges and lessons learned

ObjectivesSeroprevalence surveys provide crucial information on cumulative severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) exposure. This Slovenian nationwide population study is the first longitudinal 6-month serosurvey using probability-based samples across all age categories.MethodsEach participant supplied two blood samples: 1316 samples in April 2020 (first round) and 1211 in October/November 2020 (second round). The first-round sera were tested using Euroimmun Anti-SARS-CoV-2 ELISA IgG (ELISA) and, because of uncertain estimates, were retested using Elecsys Anti-SARS-CoV-2 (Elecsys-N) and Elecsys Anti-SARS-CoV-2 S (Elecsys-S). The second-round sera were concomitantly tested using Elecsys-N/Elecsys-S.ResultsThe populations of both rounds matched the overall population (n = 3000), with minor settlement type and age differences. The first-round seroprevalence corrected for the ELISA manufacturer's specificity was 2.78% (95% highest density interval [HDI] 1.81%–3.80%), corrected using pooled ELISA specificity calculated from published data 0.93% (95% CI 0.00%–2.65%), and based on Elecsys-N/Elecsys-S results 0.87% (95% HDI 0.40%–1.38%). The second-round unadjusted lower limit of seroprevalence on 11 November 2020 was 4.06% (95% HDI 2.97%–5.16%) and on 3 October 2020, unadjusted upper limit was 4.29% (95% HDI 3.18%–5.47%).ConclusionsSARS-CoV-2 seroprevalence in Slovenia increased four-fold from late April to October/November 2020, mainly due to a devastating second wave. Significant logistic/methodological challenges accompanied both rounds. The main lessons learned were a need for caution when relying on manufacturer-generated assay evaluation data, the importance of multiple manufacturer-independent assay performance assessments, the need for concomitant use of highly-specific serological assays targeting different SARS-CoV-2 proteins in serosurveys conducted in low-prevalence settings or during epidemic exponential growth and the usefulness of a Bayesian approach for overcoming complex methodological challenges.     

Pooling Urine Samples for Ligase Chain Reaction Screening for Genital Chlamydia trachomatis Infection in Asymptomatic Women

The accuracy of pooling urine samples for the detection of genital Chlamydia trachomatis infection by ligase chain reaction (LCR) was examined. A model was also developed to determine the number of samples to be pooled for optimal cost savings at various population prevalences. Estimated costs included technician time, laboratory consumables, and assay costs of testing pooled samples and retesting individual specimens from presumptive positive pools. Estimation of population prevalence based on the pooled LCR results was also applied. After individual urine specimens were processed, 568 specimens were pooled by 4 into 142 pools and another 520 specimens were pooled by 10 into 52 pools. For comparison, all 1,088 urine specimens were tested individually. The sample-to-cut-off ratio was lowered from 1.0 to 0.2 for pooled samples, after a pilot study which tested 148 samples pooled by 4 was conducted. The pooling algorithm was 100% (48 of 48) sensitive when samples were pooled by 4 and 98.4% (61 of 62) sensitive when samples were pooled by 10. Although 2.0% (2 of 99) of the negative pools of 4 and 7.1% (1 of 14) of the negative pools of 10 tested presumptive positive, all samples in these presumptive-positive pools were negative when retested individually, making the pooling algorithm 100% specific. In a population with 8% genital C. trachomatis prevalence, pooling by four would reduce costs by 39%. The model demonstrated that with a lower prevalence of 2%, pooling eight samples would reduce costs by 59%. Pooling urine samples for detection of C. trachomatis by LCR is sensitive, specific, and cost saving compared to testing individual samples.     

Use of pooled urine samples and automated DNA isolation to achieve improved sensitivity and cost-effectiveness of large-scale testing for Chlamydia trachomatis in pregnant women

The success of large-scale screening for Chlamydia trachomatis depends on the availability of noninvasive samples, low costs, and high-quality testing. To evaluate C. trachomatis testing with pregnant women, first-void urine specimens from 750 consecutive asymptomatic pregnant women from the Rotterdam area (The Netherlands) were collected. Initially, we investigated the performance of three different DNA isolation methods with 350 of these urines and 70 pools of 5 of the same subset of urine samples. The routinely used COBAS AMPLICOR test was compared to the COBAS AMPLICOR test with prior DNA isolation by use of the MagNA Pure large-volume kit and the MagNA Pure bacterial DNA isolation kit. The latter combination provided the best DNA test for pooled urines, with a sensitivity twice that of the other methods. Next, using all 750 urines, the COBAS AMPLICOR performance for individual testing was compared to pooled testing with the standard COBAS AMPLICOR procedure and subsequently to pooled testing with COBAS AMPLICOR in combination with the MagNA Pure bacterial DNA isolation kit. The sensitivity of COBAS AMPLICOR was 65% on individual and 42% on pooled urines but improved to 92% on pooled urines with the MagNA Pure bacterial DNA isolation kit, making this combination the best screening method. The C. trachomatis prevalence in this population appeared to be 6.4%. Additionally, the cost of the combined MagNA Pure bacterial DNA isolation kit and COBAS AMPLICOR method on pooled urines was only 56% of the cost of the standard COBAS AMPLICOR test applied to individual urines. Costs per positive case detected in the combined method were 39% of standard costs.     

Evaluation of pooled rapid HIV antibody screening of patients admitted to a San Diego Hospital

Current HIV screening guidelines in the United States recommend expanding the scope of HIV screening to include routine screening in health care settings; however, this will require increased resources. Since testing of pooled samples can decrease costs, the test characteristics of pooled rapid antibody testing were determined and optimal pool sizes were estimated for populations with HIV prevalence ranging from 0.25% to 10%. Based on these results, pooled testing methods were evaluated for screening patients admitted to hospital in San Diego, California. Evaluation of pooled antibody testing on samples collected from individuals with known HIV infection found only a modest reduction in sensitivity. These false negative results were only found among samples with very low optical density readings (<0.125 by the ADVIA Centaur? HIV assay). These readings are considered as HIV negative by the ADVIA Centaur? HIV assay, and therefore likely correspond to samples collected during acute infection. Further evaluation of pooled testing of samples collected from individuals during recent infection, found that mini-pool testing of five samples detected HIV antibody in 86% of samples taken within 60 days of the initial infection and 92% of samples taken within 90 days of the initial infection. Based on estimations of optimal pool sizes for low prevalence populations, it was decided to evaluate mini-pools consisting of 10 samples to screen the study's hospitalized patients. During this evaluation, the HIV prevalence among hospitalized patients was 0.8%, and the 10 sample mini-pool testing had 100% sensitivity and specificity. Additionally, pooled testing resulted in an 84.5% reduction in the number of rapid HIV antibody tests needed, as compared to testing each sample individually. Even when incorporating the increased costs of technician time, mini-pooled tested would have resulted in a net savings of 8760 USD for the 523 samples tested in the study. Taken together, these results indicate that pooled rapid antibody testing may reduce substantially the costs for HIV screening in low prevalence populations without a loss in accuracy.     

Blind comparison of Abbott and Dupont HIV antigen ELISA tests for detecting antigenaemia in asymptomatic human immunodeficiency virus antibody positive homosexual men.

Three hundred and ninety eight serum samples from 270 human immunodeficiency virus (HIV) antibody positive asymptomatic homosexual men were tested in the Abbott and Dupont HIV antigen ELISA tests. In the Abbott test 62 (16%) of the sera were positive, according to the manufacturer's instructions, compared with 55 (14%) in the Dupont test. Twenty six sera were positive with the Abbott test but negative with the Dupont test and 19 sera were positive only by the Dupont test. Only 36 (9%) of the sera were positive in both tests. The Abbott confirmatory neutralisation test gave excellent agreement with the initial Abbott HIV antigen ELISA test; the Dupont confirmatory test was only in agreement with the initial positive Dupont antigen ELISA test in one third of the sera tested. Although the overall sensitivity of each of the two commercial assays tested was similar, the Abbott method may be preferable for clinical purposes if confirmation of an initial ELISA positive test result by neutralisation assay is required.     

Seroprevalence of HHV 8 antibodies among the general population and HIV positive persons in the Czech Republic.

BACKGROUND: The seroprevalence rates of herpesvirus 8 (HHV 8) antibodies were determined for the general Czech population and HIV-positive individuals. OBJECTIVES: Six hundred and sixty six serum samples from the general Czech population and 129 serum samples from HIV-positive persons were tested for the presence of antibodies to the HHV 8 lytic and latent antigens. STUDY DESIGN: HHV 8 antibodies were detected by the indirect immunofluorescence test. RESULTS: In the general Czech population, only 2.4 and 0.3% of the serum samples tested positive for antibodies against the lytic and latent HHV 8 antigens, respectively. As many as 34.9 and 10.9% HIV positive individuals had antibodies to the HHV 8 antigens, respectively. Only three of them have developed Kaposi's sarcoma (KS) to date. At the time of KS diagnosis, the three patients had antibodies to both HHV 8 antigens. HIV-positive homo/bisexuals were at significantly higher risk of acquiring HHV 8 infection compared with HIV-positive heterosexuals. The increase in HHV 8 seroprevalence was associated with progression of the HIV infection from stage A to stage B. No correlation was found between the HHV 8 seroprevalence and CD 4+T-lymphocytes counts or the HIV viral load. CONCLUSIONS: Among the general Czech population, the HHV 8 seroprevalence is as low as in the West European countries. The mean HHV 8 seroprevalence rate in HIV-positive individuals was 34.9% and was comparable with those reported in other low seroprevalence countries.     

Antibody detection using pooled sera and a solid phase system

The purpose of the study was to evaluate the feasibility of substituting the Immucor Capture-R solid phase (SP) antibody detection system for our routine donor antibody screen. Our routine procedure (RP) used a 12-drop pool of six donor sera and one drop of pooled reagent red cells, with 37 degrees C incubation and indirect antiglobulin test readings. The SP system was used according to the manufacturer's directions except that one drop of the pooled sera (rather than an individual serum) was added to each microwell. In parallel tests of 888 donor pools and of 70 stored sera containing known alloantibodies, results with SP were comparable to results with RP. After implementation of SP for donor antibody detection, 1,135 donor pools were tested. Results with SP appeared satisfactory when compared to previous records. We concluded that Immucor Capture-R system can be used for antibody detection using pooled donor sera.     

Identification of a high-risk heterosexual population for HIV prevention trials in Rio de Janeiro, Brazil. Projeto Praço Onze Study Group

The study of interventions to prevent HIV transmission requires access to populations with a high rate of HIV transmission. We estimated HIV incidence among heterosexual males and females who were seen at an HIV testing site in Rio de Janeiro, Brazil. Stored sera from individuals who visited the site between March and December 1998 were analyzed using the sensitive/less sensitive (S/LS) assay and a chart abstraction was performed. During the study period, 6353 serum samples were tested. Of those tested, 1203 were found to be HIV-seropositive or indeterminate, of which 1050 (87%) remained available for further testing. In addition, 84 serum samples, representing 63 adults, were found to produce results suggesting early HIV infection. Of these, 14 were heterosexual and female (median age, 38 years), and 19 were heterosexual and male (median age, 25 years). The estimated HIV seroincidence was 1.9 (95% confidence limits (CL), 0.9%-3.9%) and 2.8 (95% CL, 1.4%-5.3%) per 100 person-years among heterosexual women and men, respectively. A survey on willingness to participate in future placebo-controlled HIV vaccine trials in this population indicated that 54.5% and 53.9% of heterosexual women and men, respectively, indicated that they would definitely be willing to participate. We have identified a heterosexual population in Rio de Janeiro with a high rate of HIV transmission willing to participate in placebo-controlled vaccine trials. This study demonstrates the usefulness of the newly described S/LS assay, which allows one to estimate HIV incidence from single serum specimens.     

The use of simple, rapid tests to detect antibodies to human immunodeficiency virus types 1 and 2 in pooled serum specimens.

BACKGROUND: The use of pooled specimens has been proposed as a means of expanding testing for human immunodeficiency virus (HIV) antibodies in population studies and in blood screening, while reducing laboratory costs. OBJECTIVES: To develop a strategic specimen pooling method to be used with rapid HIV antibody assays to detect positive specimens and to evaluate its performance in comparison with testing with commercial EIA and WB. STUDY DESIGN: Two lateral flow rapid HIV antibody assays, Seroz*Strip HIV-1/2(1) and Determine HIV-1/2, were evaluated for their ability to detect HIV-1 antibodies in serum and/or plasma specimens pooled in sizes ranging from two to 20 following the respective manufacturers' protocols. One thousand prospectively collected specimens and 55 seroconversion specimens were prepared in pools of five for evaluation by the two rapid HIV assays. RESULTS: Optimal detection and discrimination of HIV-1 antibody-positive and HIV-1 antibody-negative specimens was observed in pool sizes of five to ten for both assays. The ability of the two rapid assays to detect HIV-1 antibody-positive samples from commercial HIV-1 seroconversion panels contained in the pools was equivalent to that of commercial enzyme immunoassays (EIAs) and Western blot (WB) to detect HIV-1 antibody in the non-pooled samples. Application of the pooling method in prospectively collected specimens yielded excellent concordance with EIA/WB results in both sensitivity (98.88% for Seroz*Strip HIV-1/2, 100% for Determine HIV-1/2) and specificity (99.56% for Seroz*Strip HIV-1/2, 99.45% for Determine HIV-1/2). CONCLUSION: Use of a pooling strategy with either assay reduced the number of tests required by almost 50% and could provide substantial cost reductions for HIV screening in settings where HIV-1 prevalence is less than 10%.