Urine antibody tests: new insights into the dynamics of HIV-1 infection.

BACKGROUND: Noninvasive methodologies provide alternatives to diagnostic blood tests and have high patient acceptance, increased safety, and reduced costs. Such tests may supplement or replace blood diagnostic assays currently in use. METHODS: Using a licensed urine-based test for antibody to HIV-1, we performed 25 991 HIV-1 urine antibody enzyme immunoassay (EIA) screening tests [confirmable by HIV-1 Western blot (WB)] on paired urine and blood specimens obtained from high- and low-risk HIV-1 subjects collected at six sites representative of the US population. RESULTS: Using HIV-1 urine EIA tests confirmed by urine Western blot, a compartmentalized immune response (urine positive/serum negative) occurred in 0.24% of a cohort of 11 896 subjects. In the same cohort, specimens that were urine negative/serum positive occurred in 0.17% of subjects. In a second study of 25 991 subjects that included 859 high-risk individuals, the false-positive urine EIA frequency (urine WB negative or indeterminate) was 1.3%. This false-positive frequency in the high-risk cohort was attributed, in part, to an IgA antibody response. We tabulated urine and serum indeterminate reactivities and examined their possible causes. Data are presented showing that antibodies from a seroindeterminate HIV-1vau group O subject were reactive in urine EIA and urine WB tests. An analysis of the HIV-1vau strain group O env nucleotide sequence disclosed a high frequency of homology with human chromosome 7q31, a fragile site implicated in many human malignancies. CONCLUSIONS: These results demonstrate the utility of urine for alternative HIV-1 antibody testing and provide new insights into the pathogenesis of HIV-1 infection and into potential application of this approach in investigation of other microbial pathogens and toxic compounds.

Increased sensitivity of HIV-1 antibody detection.

Clinical trial results from 11,344 paired urine and serum samples revealed 1,181 HIV-1-positive individuals confirmed by western blot (WB). There were 25 discrepant samples: 10 were urine enzyme immunoassay (EIA) and WB positive, serum non-reactive and serum WB negative or indeterminate, and 15 were serum EIA and WB positive, urine EIA non-reactive or urine WB negative or indeterminate. Serum samples, HIV-1 antibody WB confirmed, revealed a 99.15% sensitivity (1,171 out of 1,181); urine samples, HIV-1 antibody WB confirmed, showed a 98.73% sensitivity (1,166 out of 1,181). This study demonstrated that neither serum nor urine results alone are as sensitive for HIV-1 antibody detection as combined results of both samples.

Legionella micdadei and Legionella dumoffii monoclonal antibodies for laboratory diagnosis of Legionella infections.

Two different monoclonal antibodies directed against Legionella micdadei and L. dumoffii (Genetic Systems Corp., Seattle, Wash.) were evaluated for their specificity and ability to detect L. micdadei and L. dumoffii in human and animal clinical samples and bacterial isolates in an indirect immunofluorescence assay. All three frozen sputum samples and all three Formalin-fixed sputum and liver samples from patients with culture-documented L. micdadei pneumonia were positive when tested with the L. micdadei monoclonal antibody. A Formalin-preserved lung sample from a patient with culture-documented L. dumoffii pneumonia was positive with its homologous monoclonal antibody. No cross-staining reactions were found with either monoclonal antibody on any of 25 human sputum samples tested from patients without Legionella infections. A total of 66 Legionella strains and 56 non-Legionella strains including 22 Pseudomonas strains and 34 other bacterial strains were studied. No cross-staining reactions were found except in Staphylococcus aureus Cowan 1 ATCC 12598. The lower limit of detection in seeded sputum samples was about 7 X 10(4) cells per ml for both monoclonal antibodies. Lung and tracheal lavage specimens from L. micdadei- or L. dumoffii-infected guinea pigs showed specific staining only with their respective monoclonal antibodies. The monoclonal antibodies stained homologous bacteria slightly less intensely than did the polyclonal antisera, but the signal-to-noise ratio was considerably higher for the monoclonal antibodies. No differences in sensitivity of staining of clinical specimens or bacterial isolates were noted between the monoclonal antibodies and the polyclonal reagents for L. micdadei and L. dumoffii (Centers for Disease Control, Atlanta, Ga., and BioDx, Denville, N.J. These monoclonal antibodies ae sensitive and specific, making them good candidates for laboratory diagnostic purposes.

Comparison of cross-staining reactions by Pseudomonas spp. and fluorescein-labeled polyclonal and monoclonal antibodies directed against Legionella pneumophila.

Commercially prepared polyclonal antisera to Legionella pneumophila are known to cross-react with organisms of the genus Pseudomonas. To determine whether a commercially available monoclonal antibody reagent specific for L. pneumophila would also cross-react with pseudomonads, a two-laboratory study was undertaken to test both monoclonal and polyclonal reagents against 33 isolates of Pseudomonas spp., including 25 Pseudomonas aeruginosa, 4 P. putida, 2 P. maltophilia, 1 P. fluorescens, and 1 P. alcaligenes. Four antisera were tested; polyclonal anti-legionella antisera pools A and B (Centers for Disease Control [CDC], Atlanta, (Ga.), polyclonal 1-6 antisera (BioDx, Inc., Denville, N.J.), and a monoclonal antibody reagent produced by Genetic Systems Corp., Seattle, Wash. All reagents were labeled with fluorescein. Cross-staining reactions were found with the BioDx L. pneumophila antisera and 10 isolates of Pseudomonas. Four of these isolates demonstrated cross-staining with CDC pool A. When tested with individual serotype-specific reagents (CDC), three of four cross-reacted with L. pneumophila serotype 1 antisera; the fourth cross-reacted with serotype 3. No cross-staining reactions were noted with the monoclonal reagent and any of the pseudomonads tested, demonstrating that the Genetic Systems Corp. monoclonal reagent is the most specific of the four reagents tested.

Clinical utility of a monoclonal direct fluorescent reagent specific for Legionella pneumophila: comparative study with other reagents.

Twenty-four lower respiratory tract samples taken from patients with culture-confirmed Legionella pneumophila infection were examined with three different direct immunofluorescent antisera to L. pneumophila, as were 29 samples from similar sources taken from patients without Legionnaires disease. The reagents studied were Genetic Systems Corp. (GS) monoclonal L. pneumophila conjugate, which reacts with all known serogroups of L. pneumophila, BioDx polyvalent L. pneumophila serogroups 1 through 6 conjugate, and Centers for Disease Control polyvalent pool A L. pneumophila serogroups 1 through 4 conjugate. The specimens had been frozen at -70 degrees C for 0.5 to 5 years. Randomization was used in coding the samples, which were stained and read by an independent observer. All three conjugates correctly identified all positive and negative samples. No difference was noted among the conjugates in the absolute numbers of fluorescent L. pneumophila bacteria per sample. The GS conjugate had a much cleaner background than did the other two reagents. Mean staining intensity scores were 3.4, 3.9, and 3.7 for the GS, BioDx, and Centers for Disease Control conjugates, respectively. This study demonstrates that the diagnostic efficiency of all three conjugates is equivalent. Since the GS conjugate is easier to read, does not cross-react with non-L. pneumophila bacteria, and reacts with serogroups 1 through 10 of L. pneumophila, it appears to be preferable for use in diagnostic testing on nonhistopathologically processed specimens.

Identification of a species-specific antigen in Legionella pneumophila by a monoclonal antibody.

A species-specific antigen in Legionella pneumophila was identified by a monoclonal antibody in enzyme-linked immunosorbent and immunofluorescence assays of serogroups 1 through 8. The species-specific antigen was heat stable, and the molecular weight of the major band was 29,000 by immunoblot analysis. In direct immunofluorescence assays, the antigen was cryptic or only partly exposed on the surface of the cells but was effectively exposed by treating the cells with detergent and EDTA. The monoclonal antibody was utilized in direct immunofluorescence assays to specifically identify multiple cultures of L. pneumophila serogroups.

Induction of target antigens and conversion to susceptible phenotype of NK-cell-resistant lymphoid cell line.

Two autologous Herpesvirus papio producer lymphoid cell lines and one autologous non-producer line were compared for susceptibility to natural killer (NK) cell-mediated lysis. The non-producer cell line, 26CB-1, was more resistant to NK cell killing compared to one viral producer counterpart 13CB-1, but equally resistant when compared to another, 8CB-1. Treatment with chemical agents that affect differentiation or activate the viral cycle, including n-butyrate, IuDR, 5-azacytidine and tunicamycin, increased the susceptibility to killing of the non-producer line but had less effect on the 13CB-1 producer line. The increase in susceptibility was due to induction of new target antigens: activated 26CB-1 cells were more effective at inhibiting NK-cell-mediated lysis and were bound by more NK cells than untreated control cells. The expression of NK target structures may be related to the differentiated state rather than to the viral production status to target cells.

Phylogeny of NK cell reactivity against human and nonhuman primate lymphoblastoid cell lines: evolving and conserved target antigens.

Both human and nonhuman primate natural killer (NK) cells display little or no killing against allogeneic B lymphoblastoid cell lines. However, the same B cell lines are killed in baboon-human (B alpha H) or human-baboon (H alpha B) xenogeneic combinations. Competition-inhibition experiments indicate that the xenogeneic determinants recognized by NK cells are found principally if not exclusively on B rather than T target cells. Cell lines from closely related chimpanzee or orang-utan species can block some killing of human target cells, but lines from more distantly related species including gibbon, macaque, baboon, and marmoset do not inhibit cytotoxicity. This suggests that some NK target structures are susceptible to evolutionary change. Gibbon or marmoset lines infected with Epstein Barr virus (EBV) do not block killing, suggesting that host rather than viral determinants are being recognized. In contrast to the foregoing pattern, 2 cell lines derived from the same baboon differed in susceptibility to NK lysis irrespective of the effector cell species. The viral producer line 13CB-1 was more susceptible to lysis than its viral nonproducer partner 26CB-1. Thus, some NK target antigens may be highly conserved whereas others evolve with the species.

Monocyte-reactive antibodies in patients with systemic lupus erythematosus.

Cold-reactive antibodies cytotoxic for peripheral monocytes from more than half of normal donors were found in the sera of 2 of 25 patients with systemic lupus erythematosus (SLE) and 1 of 26 with rheumatoid arthritis (RA), and they were absent in 25 normal sera. In contrast, lymphocytotoxic activity for T or B lymphocytes was found in over half of the lupus sera. The antibodies to monocytes were primarily IgM and exhibited varying specificities. Some of the antibodies were directed against antigenic determinants common to monocytes, T and B cells, or against determinants shared between monocytes and one lymphocyte type. One serum possessed a high titer of antibodies that were specific for monocytes. The clinical significance of antimonocyte antibodies remains to be established.