P24 Reactivity Research Articles

False-positive human immunodeficiency virus enzyme immunoassay results in pregnant women.

ObjectiveExamine whether false-positive HIV enzyme immunoassay (EIA) test results occur more frequently among pregnant women than among women who are not pregnant and men (others).DesignTo obtain a large number of pregnant women and others tested for HIV, we identified specimens tested at a national laboratory using Genetic Systems HIV-1/HIV-2 Plus O EIA from July 2007 to June 2008.MethodsSpecimens with EIA repeatedly reactive and Western blot-negative or indeterminate results were considered EIA false-positive. We compared the false-positive rate among uninfected pregnant women and others, adjusting for HIV prevalence. Among all reactive EIAs, we evaluated the proportion of false-positives, positive predictive value (PPV), and Western blot bands among indeterminates, by pregnancy status.ResultsHIV prevalence was 0.06% among 921,438 pregnant women and 1.34% among 1,103,961 others. The false-positive rate was lower for pregnant women than others (0.14% vs. 0.21%, odds ratio 0.65 [95% confidence interval 0.61, 0.70]). Pregnant women with reactive EIAs were more likely than others (p<0.01) to have Western blot-negative (52.9% vs. 9.8%) and indeterminate results (17.0% vs. 3.7%) and lower PPV (30% vs. 87%). The p24 band was detected more often among pregnant women (p<0.01).ConclusionsFalse-positive HIV EIA results were rare and occurred less frequently among pregnant women than others. Pregnant women with reactive EIAs were more likely to have negative and indeterminate Western blot results due to lower HIV prevalence and higher p24 reactivity, respectively. Indeterminate results may complicate clinical management during pregnancy. Alternative methods are needed to rule out infection in persons with reactive EIAs from low prevalence populations.

P096 HIV p24 reactivity in Hungarian blood donors without evidence of HIV infection

P096 HIV p24 reactivity in Hungarian blood donors without evidence of HIV infection

Macrophage levels in large cell lymphoma are associated with survival

8097 Background: Recent studies of follicular lymphoma have suggested that the presence of high levels of tumor associated macrophages (MO) conferred a poor prognosis. The current study evaluated large cell lymphomas for the presence of MOs and “proliferative MOs” (promacs), in HIV+ and HIV- LCL. Methods: Formalin-fixed and paraffin-embedded tissues from patients with LCL (and controls) were obtained from the ACSR (URL: http://acsr.ucsf.edu ) and evaluated for expression of CD68 (MO), PCNA (proliferation) and HIVp24 via the use of 0.6mm section tissue microarrays (TMA). Survival data was also evaluated. Results: 64 cases of LCL were evaluated, 31 of which were HIV+ (33 cases were HIV-). HIV p24 staining was seen with 13 of the 31 HIV+ cases(42%) , with staining only in MOs. Promacs (CD68+/PCNA+MOs) were more frequent in cases of HIV+ LCL (median of 18) and HIV- LCL (median of 5), than normal lymph nodes and reactive tonsillar tissue (p&lt;.001). In both HIV+ and HIV- cases of LCL, there was a trend towards shorter survival (p&lt;0.07) with increased numbers of MOs and promacs; p24 reactivity was unrelated to survival. Conclusions: In this preliminary study of MOs in LCL, MO number per TMA core was significantly correlated with promac number in all cases of LCL (p&lt;.001). HIV p24 localized to MOs in 42% of HIV+ LCL cases and was unrelated to survival. Increased promac numbers in cases of LCL correlated with decreased survival in all cases of LCL, but cases of HIV+LCL tended to demonstrate shorter survival than in cases of HIV- LCL. Of the latter group, higher numbers of macrophages (for example, greater than 46 macrophages/core) portend a worse prognosis, but this finding was not statistically significant in this preliminary study. No significant financial relationships to disclose.

Optimized virus disruption improves detection of HIV-1 p24 in particles and uncovers a p24 reactivity in patients with undetectable HIV-1 RNA under long-term HAART

HIV-1 p24 antigen (p24) measurement by signal amplification-boosted ELISA of heat-denatured plasma is being evaluated as an alternative to HIV-1 RNA quantitation in resource-poor settings. Some observations suggested that virion-associated p24 is suboptimally detected using Triton X-100-based virus dissociation buffer (kit buffer). A new reagent (SNCR buffer) containing both denaturing and non-denaturing detergents was therefore developed and evaluated. The SNCR buffer increased the measured p24 concentration about 1.5- to 3-fold in HIV-negative plasma reconstituted with purified HIV-1 particles, while not increasing the background. Among 127 samples of HIV-1-positive patients with moderate to high concentrations of HIV-1 RNA the increase was about threefold across the entire concentration range (P < 0.0001). Specificity before neutralization among prospectively tested clinical samples ruled HIV-negative was 828 of 845 (98.0%) for the SNCR buffer and 464 of 479 (96.9%) for kit buffer. Specificity after confirmatory neutralization of reactive samples or a follow-up test was 100% with either buffer. Surprisingly, the SNCR buffer revealed a p24 reactivity in 115 of 187 samples (61.5%) from adult patients exhibiting undetectable HIV-1 RNA below 5 copies/ml for a duration of 6-30 months under HAART (3.7% with kit buffer). The rate of p24 reactivity in these patients did not decrease with duration of HAART. In conclusion, the SNCR buffer improves the detection of particle-associated HIV-1 p24, thereby increasing the measured p24 concentration in samples with medium to high HIV-1 RNA. It also uncovers the presence of a p24 reactivity, whose identity remains to be determined, in a significant fraction of samples with undetectable HIV-1 RNA under long-term HAART.

Indeterminate human immunodeficiency virus Western blot profiles in ethiopians with discordant screening-assay results.

The Western blot (WB) assay is the most widely accepted confirmatory assay for the detection of antibodies to human immunodeficiency virus type 1 (HIV-1). However, indeterminate WB reactivity to HIV-1 proteins may occur in individuals who do not appear to be infected with HIV. The profiles of WB reactivity among Ethiopians are hardly known. Here, we describe the profiles of indeterminate WB reactivity in Ethiopians with discordant screening assays. Between 1996 and 2000, a total of 12,124 specimens were tested for HIV-1 antibodies. Overall, 1,437 (11.9%) were positive for HIV-1 antibody. Ninety-one ( approximately 0.8%) gave equivocal results because of discordant results among the various screening assays and indeterminate WB profiles by the American Red Cross (ARC) criteria. Most (30.4%) of these indeterminate WB results were due to p24 reactivity. However, 12 samples (13.2%) displayed reactivity to p24 and gp41 or to p24 and gp120/160 proteins (positive by Centers for Disease Control and Prevention [CDC] criteria). Only two samples (2.2%) were reactive to both env glycoproteins gp41 and gp120/160 (positive by the World Health Organization [WHO] criteria). Of 31 WB assays initially indeterminate by the ARC criteria and with follow-up samples, 29 (93.5%) became negative when retested subsequently while 2 (6.5%) remained indeterminate for more than a year and were thus considered negative. Using CDC and WHO criteria, 6 (19.4%) and 2 (6.5%), respectively, of these WB assays would have been considered falsely positive. In addition, 17 indeterminate samples were negative when assessed by a nucleic acid-based amplification assay for HIV-1 viremia. In general, there was 97.8% concordance between the ARC and WHO criteria and 85.7% concordance between the ARC and CDC criteria for an indeterminate WB result. The ARC criteria best met the specified objectives for diagnosis in our setting.

Detection of HIV‐1 RNA in Two Consecutive Blood Donations Screened Negative for HIV Antibodies

Abstract Background and Objectives: An unusual serological pattern of HIV‐1 seroconversion in a blood donor is described. The seroconversion panel was used to investigate the sensitivity of existing screening assays. Materials and Methods: A volunteer blood donor who had given blood 79 times was diagnosed anti‐HIV‐1‐antibody‐positive. The heteroduplex mobility assay identified a subtype B HIV‐1 strain. The frozen plasmas from the last four blood donations had been kept at −30°C. They were thawed and aliquoted for subsequent testing. Results: The last two blood donations contained HIV‐1 RNA, 2,800 copies/ml (October 26) and 170 copies/ml (November 23). Weak anti‐p24 antibodies were detected by Western blot in the October 26 sample, and a clear p24 reactivity along with a faint gp 160 reactivity was oberved on November 23. HIV p24 antigen was undetectable in both samples. Out of 13 screening assays, only 6 gave positive results on the November sample and 7 negative results which were obtained by 1 competitive enzyme immunoassay (EIA) and 6 of the 9 sandwich EIAs. Conclusion: Most sandwich EIAs gave prolonged false‐negative results in the present case. p24 antigen testing was negative and would not have reduced the risk of HIV transmission.

Humoral Aspects of Anti-HIV Immune Responses in Zairians with AIDS: Lower Antigenemia Does Not Correlate with Immune Complex Levels

Serological patterns of anti-HIV immune responses of 150 HIV-infected (65 asymptomatic, 19 ARC, 66 AIDS) and 150 HIV-negative healthy Zairians were studied to determine the clinical significance of p24 antigen, and anti-p24 antibody, particularly in relation to p24 relative binding capacity (RBC) and circulating immune complexes (CICs). Levels of p24 antigen, anti-p24 antibody titers, and p24 RBC were evaluated by means of enzyme-linked immunosorbent assay (ELISA). Circulating immune complexes were measured by C1q-binding assay. Human immunodeficiency virus CICs were detected by polyethylene glycol (PEG) precipitation followed by 6 M guanidinium lysis, ELISA, Western blot, or radioimmunoprecipitation of the lysed precipitates. Immunoglobulin levels for IgG, IgM, IgA, and beta 2-microglobulin (beta 2-M) were quantified in all study participants by laser nephelometry and ELISA. All immunoglobulin levels were significantly elevated among HIV-positive vs. HIV-negative individuals. Immunoglobulin levels correlated well with disease progression among infected patients. Similarly, beta 2-M levels were significantly higher in HIV-positive vs. HIV-negative individuals and correlated well with progression to AIDS. Free p24 antigen in serum was detected in 1.33% of all patients. However, p24 reactivity increased to 6% (9 of 150 cases) after PEG precipitation and CIC dissociation. There was a good correlation between p24 reactivity and disease development. High titers of anti-p24 antibody (> 44,100) occurred in at least 80% of all patients, and did not correlate with disease stage. Similarly, more than 60% of patients had high levels of p24 RBC.(ABSTRACT TRUNCATED AT 250 WORDS)

Identification of Regions of HIV-1 p24 Reactive with Sera Which Give "Indeterminate" Results in Electrophoretic Immunoblots with the Help of Long Synthetic Peptides

We analyzed nine sera from persons unlikely to be HIV infected which had an IgG reactivity directed against HIV-1 p24, and in two cases also to its precursor p55, but to no other HIV proteins, nor to proteins of the H9 host cell, in electrophoretic immunoblots (EIB). These sera are also referred to as having an indeterminate HIV EIB pattern or as HIV antibody false positive sera. Seven of nine sera reacted with longer (61-77 amino acids) and none with shorter (17-25 amino acids) p24-derived peptides in enzyme immunoassays (EIAs). This is compatible with a conformational (discontinuous) nature of the epitopes involved in many false positive HIV-1 p24 antibody reactions. Four sera reacted with an N-terminal, one with an internal, and two with a C-terminal fragment. Each of the seven sera thus only reacted with one of the long p24 peptides. The specificity and singularity of the reaction was further demonstrated by competition and/or absorption experiments with synthetic peptides. In contrast, 18 of 20 confirmed HIV-1+ sera with p24 reactivity in EIB reacted with at least one and often several of the longer peptides, most frequently the C-terminal one. Thus, the distribution of peptide reactivity of true HIV-1 antibody-positive sera was different from that of the falsely reactive sera. According to two of several explanations, these antibodies may have arisen because of (1) molecular mimicry by chance or by functional selection, (2) immunization by activation, noninfectious exposure, or infection involving non-HIV endogenous or exogenous retroviral antigens. The latter gains some support from our finding of antibody reactions with capsid proteins of the simian viruses, simian sarcoma-associated virus (SSAV), and Mason-Pfizer monkey retrovirus in some of the p24 +/- p55 reactive sera.

Characterization of antibody reactivity to human T-cell lymphotropic virus types I and II using immunoblot and radioimmunoprecipitation assays

We have characterized the immunoreactivity to human T-cell lymphotropic virus type I (HTLV-I) among 26,983 persons of various seroprevalence groups by using enzyme immunoassay, immunoblot (IB), and radioimmunoprecipitation assays (RIPA) in accordance with Public Health Service recommended guidelines for the interpretation of serologic test results for HTLV-I infection. IB-indeterminate serum specimens (n = 178) were reactive to HTLV-I gag proteins, and no serum contained only env reactivity. Overall, RIPA resolved 40% of IB-indeterminate serum samples; however, the probability that RIPA would confirm IB-indeterminate samples depended on the seroprevalence of the population tested. HTLV-I gag p19-only reactivity on IB was not a reliable marker of HTLV-I infection, while gag p24 reactivity on IB was clearly associated with positive seroreactive specimens. IB and RIPA tests did not clearly distinguish between HTLV-I and HTLV-II seroreactivities. These data emphasize that patterns of immunoreactivity to HTLV-I antigens are dependent upon the seroprevalence of the risk groups tested. In addition, RIPA detected antibodies to env proteins present in low titer in a substantial number of IB gag-only reactive sera and resolved the HTLV-I antibody status of these sera.

Investigation of atypical western blot (immunoblot) reactivity involving core proteins of human immunodeficiency virus type

Serum specimens which originally exhibited a narrow (indeterminate) 24-kilodalton core protein (p24) or p24/p55 pattern of reactivity with human immunodeficiency virus (HIV) in the Western blot (immunoblot) test were studied to gather information on antibody specificity. A total of 12 specimens were initially reevaluated with an indirect immunofluorescence assay (IFA), three enzyme-linked immunosorbent assays (ELISAs), and Western blot analyses. Five of the specimens were IFA positive and contained anti-gp160/gp120 antibodies which were observed only when an HIV Western blot antigen rich in gp160 and gp120 was used. The remaining seven serum specimens were nonreactive by IFA and showed variable reactivity in HIV antibody ELISAs. The specimens did not cross-react with core antigens for human T-cell leukemia virus types 1 and 2 or contain detectable levels of HIV p24 antigen. The p24/p55 reactivity of six of the seven indeterminate specimens could be reduced or eliminated by preincubating the specimens with disrupted, HIV-infected H9 cells but not with uninfected H9 cells. The six specimens also exhibited discernible reactivity with recombinant HIV p24 antigen. When an additional 23 indeterminate specimens were assayed, all of the serum specimens were nonreactive by IFA while 65% (15 of 23) showed various degrees of reactivity with the recombinant p24 protein. There was no indication that any of the HIV core antibody reactivity was caused by HIV infection. Indeterminate results for five patients with specific p24 reactivity, who were retested after a period of weeks or months, remained indeterminate for HIV antibody with no significant change in ELISA or Western blot reactivity.

HIV‐Antibody Testing and False‐Positive p24 Reactivity

HIV‐Antibody Testing and False‐Positive p24 Reactivity

Follow-up testing and notification of anti-HIV Western blot atypical (indeterminant) donors.

We conducted anti-HIV testing on follow-up samples obtained at a mean interval of 20 weeks from 150 blood donors who had previously tested anti-HIV ELISA positive and Western blot atypical. Of 93 donors who demonstrated reactivity to HIV core protein p24, 4 progressed to positive Western blots. Most of the remaining donors showed a persistent p24 reactivity on Western blot and had no risk factors for HIV infection. Immunofluorescence testing of the initial sample from 93 donors could not definitively separate seroconverters from those with persistent p24 reactivity. Of 57 donors with p18 reactivity, none were positive on follow-up anti-HIV testing. Our findings suggest policies and strategies for notifying donors of atypical anti-HIV Western blot results.

Sequential changes in antibody levels to the env and gag antigens in human immunodeficiency virus infected subjects.

Sera from 51 HTLV-III (human immunodeficiency virus, HIV)-antibody positive subjects consisting of 21 asymptomatic individuals and 15 ARC and 15 AIDS patients were analyzed for their serological profiles toward the viral antigens. One of the asymptomatic subjects only showed a p24 reactivity in the immunoblot, but antibodies to the env antigens were clearly identified by immunoprecipitation of viral antigens (RIP) followed by SDS-polyacrylamide gel electrophoresis. RIP patterns of different subjects and even different bleeds from the same subjects showed a varying reactivity to the gag antigens whereas the reactivity towards the env antigens appeared to be generally stable. RIP analysis of sequential sera of virus-infected individuals indicated a pattern consistent with an initial steady rise of antibody reactivities to the gag antigens relative to the reactivities to the envelope antigens. These reactivities reached a plateau and then slowly declined. While all sera tested had antibodies to the envelope antigens gp160, gp120 and gp41, 86% of the asymptomatic subjects, 67% of the ARC patients and only 33% of the AIDS patients had antibodies to the gag proteins p24 and pr53gag.