Biomedical subjects

J Schüpbach

Publications and source records attributed to J Schüpbach.

At least 37 records · Page 2Linked to original sources

PubMed1998

The role of in vitro-induced lymphocyte apoptosis in feline immunodeficiency virus infection: correlation with different markers of disease progression.

Human immunodeficiency virus infection is characterized by a progressive decline in the number of peripheral blood CD4(+) T lymphocytes, which finally leads to AIDS. This T-cell decline correlates with the degree of in vitro-induced lymphocyte apoptosis. However, such a correlation has not yet been described in feline AIDS, caused by feline immunodeficiency virus (FIV) infection. We therefore investigated the intensity of in vitro-induced apoptosis in peripheral blood lymphocytes from cats experimentally infected with a Swiss isolate of FIV for 1 year and for 6 years and from a number of long-term FIV-infected cats which were coinfected with feline leukemia virus. Purified peripheral blood lymphocytes were either cultured overnight under nonstimulating conditions or stimulated with phytohemagglutinin and interleukin-2 for 60 h. Under stimulating conditions, the isolates from the infected cats showed significantly higher relative counts of apoptotic cells than did those from noninfected controls (1-year-infected cats, P = 0.01; 6-year-infected cats, P = 0.006). The frequency of in vitro-induced apoptosis was inversely correlated with the CD4(+) cell count (P = 0. 002), bright CD8(+) cell count (P = 0.009), and CD4/CD8 ratio (P = 0. 01) and directly correlated with the percentage of bright major histocompatibility complex class II-positive peripheral blood lymphocytes (P = 0.004). However, we found no correlation between in vitro-induced apoptosis and the viral load in serum samples. Coinfection with feline leukemia virus enhanced the degree of in vitro-induced apoptosis compared with that in FIV monoinfected cats. We concluded that the degree of in vitro-induced apoptosis was closely related to FIV-mediated T-cell depletion and lymphocyte activation and could be used as an additional marker for disease progression in FIV infection.

Animals↗

PubMed1997

A human endogenous retroviral superantigen as candidate autoimmune gene in type I diabetes.

Microbial superantigens (SAGs) have been implicated in the pathogenesis of human autoimmune diseases. Preferential expansion of the Vveta7 T cell receptor positive T cell subset in patients suffering from acute-onset type I diabetes has indicated the presence of a surface membrane-bound SAG. Here, we have isolated a novel mouse mammary tumor virus-related human endogenous retrovirus. We further show that the N-terminal moiety of the envelope gene encodes an MHC class II-dependent SAG. We propose that expression of this SAG, induced in extrapancreatic and professional antigen-presenting cells, leads to beta-cell destruction via the systemic activation of autoreactive T cells. The SAG encoded by this novel retrovirus thus constitutes a candidate autoimmune gene in type I diabetes.

Adolescent↗

PubMed1997

Simple monitoring of antiretroviral therapy with a signal-amplification-boosted HIV-1 p24 antigen assay with heat-denatured plasma.

OBJECTIVE: Virus load determination has become indispensable for the management of HIV patients, but depends on expensive assays of a low throughput. We evaluated whether a highly improved HIV-1 p24 antigen detection procedure which involves heat-mediated immune complex dissociation and signal-amplification-boosted enzyme-linked immunosorbent assay (ELISA) was suitable for antiretroviral treatment monitoring. DESIGN AND METHODS: Virus load in plasma was determined for 127 plasma samples taken at 0, 2, 6, 12, 18, 24, 30 and 36 weeks from 23 patients with CD4+ T cells < 50 x 10(6)/l who received indinavir 800 mg three times daily in addition to prior antiretroviral treatment. Tests included polymerase chain reaction (PCR) for viral RNA, measured prospectively with the Roche Amplicor kit, and retrospective batch testing of heat-denatured samples for p24 antigen by the DuPont HIV-1 p24 Core Profile ELISA linked with a tyramide signal amplification step. Particle-associated reverse transcriptase (RT) by the product-enhanced RT (PERT) assay was determined as an independent third-opinion viral load marker. RESULTS: p24 antigen was detected as sensitively as viral RNA. Overall detection during a median observation time of 25 weeks (range, 0-39) amounted to 75.6% for antigen and 73.6% for RNA. The antigen detection limit was 0.2 pg/ml. Antigen was detectable in all 23 baseline samples, whereas RNA was undetectable in one. Antigen and RNA levels in 79 samples positive for both markers correlated with r = 0.714 (P < 0.0001). Average changes in levels of p24 antigen and RNA at eight timepoints correlated with r = 0.982 (P < 0.0001). In individual patients, the two parameters behaved similarly, and in certain cases virtually identically. RT activity was measurable in all samples. CONCLUSIONS: The performance of this antigen detection procedure is comparable to RNA PCR, thus providing a simple, high throughput alternative in monitoring the efficacy of antiretroviral treatment.

Adult↗

PubMed1997

Reverse transcriptase activity in chicken embryo fibroblast culture supernatants is associated with particles containing endogenous avian retrovirus EAV-0 RNA.

We have recently shown that live attenuated virus vaccines produced on chicken-derived cells contain low levels of particle-associated reverse transcriptase (RT). In both virus and corresponding control harvests produced on chicken embryo fibroblasts, these activities were present at significantly higher concentrations than in the vaccines. In order to identify the putative retrovirus sequence responsible for this activity, a novel method for the selective PCR amplification of particle-associated retrovirus RNA that uses DNA primers complementary to the primer binding sites of the known exogenous retroviruses in combination with an anchor primer was applied. A product of the endogenous avian retrovirus family EAV-0, termed EAV-0(B1), was reproducibly generated with a tRNA(Trp)-derived primer from the RT peak fraction of a sucrose density gradient run with a harvest of a live attenuated measles vaccine. In contrast, no products were detected with primers derived from tRNA(Pro), tRNA(Lys)1,2 or tRNA(Lys)3. In the same fraction, genomic RNA of EAV-0(B1) was demonstrated by long PCR. Analysis of several sucrose density gradients from different harvests of various manufacturers demonstrated accumulation of, and colocalization with, RT activity for the EAV-0(B1) RNA but not for a chicken cellular mRNA. Synthesis of cDNA from EAV-0(B1) RNA was shown by endogenous RT reaction. Furthermore, complexes of naturally primed EAV-0(B1) RNA with RT were demonstrated. Taken together, these data strongly suggest that EAV-0 is able to produce virus-like particles with an active RT.

Alpharetrovirus↗

PubMed1996

Sensitive detection and quantification of particle-associated reverse transcriptase in plasma of HIV-1-infected individuals by the product-enhanced reverse transcriptase (PERT) assay.

Tests for the enzyme reverse transcriptase (RT) should permit the detection of all infectious retroviruses, provided that these are present as extracellular particles. The capability of a new procedure, named product-enhanced reverse transcriptase (PERT) assay, to detect HIV-1 in fresh human plasma was compared with that of the polymerase chain reaction (PCR) for viral RNA. Both procedures had identical dilution endpoints corresponding to 10(2) particles/ml. All 30 samples from HIV-1 positive patients at different stages contained RT activity whose level was significantly correlated with viral RNA and corresponded to 553-417,000 particles/ml. In HIV-1 low titer performance and seroconversion panels, the PERT assay detected more positives than PCR for viral RNA. Three of 160 blood donors exhibited elevated RT activity, indicating a prevalence of 1.9% (95% CI 0.4-5.3%). One positive donor, with laboratory parameters suggesting a mild chronic liver impairment, exhibited RT activity comparable to that of HIV positives, but was consistently negative by various tests for hepatitis viruses, cytomegalovirus, the HIVs and HTLVs. The results suggest that the PERT assay is more sensitive for detection of HIV-1 contamination of plasma than RNA PCR. However, it is not affected adversely by viral sequence variability, and may therefore, also detect HIV-1 subtype O, and additional retroviruses as yet undetectable by PCR.

Base Sequence↗

PubMed1996

Detectionof reverse transcriptase activity in live attenuated virus vaccines.

BACKGROUND: Safety considerations require that biological products for human use are free from any agent that might pose a potential health hazard. One method to detect the presence of retroviral particles is the reverse transcriptase (RT) assay. This assay is capable of detecting all infectious retrovirus particles, irrespective of genome or protein composition. Recently, a family of ultrasensitive RT tests, named product-enhanced reverse transcriptase (PERT) assays, has been designed with a detection limit that is 10(6) - 10(7) times lower than that of conventional RT tests. OBJECTIVES: To investigate with the PERT assay whether RT activity is detectable in live attenuated virus vaccines and to characterize eventual RT activities. STUDY DESIGN: A total of 12 different monovalent and one trivalent virus vaccines containing live attenuated viruses were tested for RT activity with the PERT assay and a conventional RT test. RT activities were investigated with respect to their susceptibility to RT inhibitors, association with physical particles, and their possible origin. RESULTS: One trivalent and five different monovalent vaccines contained RT activity when tested with the PERT assay, but were negative in a conventional RT assay. All lots tested of these vaccines showed RT activity. The activity in all vaccines was sensitive to AZT-triphosphate and ddTTP and at least part of it was associated with particles. Mg(2+)-dependent RT activity banded at a density of 1.14 g/ml. All positive vaccines were produced using chicken cells. CONCLUSIONS: The data indicate the systematic presence of partially particle-associated retroviral reverse transcriptase in attenuated live virus vaccines that are produced in chicken-derived cells. The identification and further characterization of these particles, as well as the elucidation of possible interactions with the human organism are imperative goals despite the fact that these vaccines have been safely used for many years.

Journal Article↗

PubMed1996

Licencing of diagnostic HIV kits-to keep a sensible standpoint in an area of permanent revolution.

BACKGROUND: In vitro diagnostic medical devices or 'test kits' are important tools for diagnosing illnesses and monitoring the patients' state of health or the effects of medical treatment. It is in the public interest that commercial kits are of high quality. OBJECTIVES: To review the current situation of kit evaluation in Europe and to establish practical and sensible criteria for kit evaluation in general, based on the Swiss experience with the licencing of kits for the detection of human immunodeficiency virus (HIV) infection. Kit evaluation should provide the user with a selection of sufficiently good products. In addition, it should be simple and not require rare patient materials. Also of importance is that all manufacturers are treated equally. METHODS: The evaluation criteria depend on the legal situation, the intended use of the test, the state of the art and the graveness of false-negative or -positive results. In addition to diagnostic sensitivity and specificity, other important considerations include the quality control by the manufacturer, kit presentation, intra- and inter-lot reproducibility and how a test fits into a country's adopted testing system. RESULTS: The above goals can be achieved by setting minimal performance standards, preferably in the form of a lower limit of the confidence interval for the sensitivity/specificity that has been observed in the test's field evaluation. The better the test, the fewer samples have to be studied. For tests with no false results observed during field trials, the minimum sample number necessary for demonstration of a sensitivity or specificity >/= 99% is about 400, while it is 700 for a >/= 99.5% demonstration. Further insight into sensitivity is provided by results from well described and frequently used commercial seroconversion panels. This represents a dynamic tool that readily identifies insensitive kits that have become technically obsolete. Specificity assessment should not only involve blood donors, but also appropriate clinical controls. It is clear that the test design has an important bearing with respect to sensitivity and specificity. There are inherent advantages and disadvantages of indirect binding assays, capture assays and double antigen sandwich assays. Positive or negative results with these different test formats may not carry exactly the same message. CONCLUSION: Efficient kit evaluation does not require tens of thousands of samples but can be performed with relatively little disbursement on materials and manpower.

Journal Article↗

PubMed1996

Isolation of an in vitro transmissible agent with reverse transcriptase activity from a blood donor with a borderline-positive HIV-1 serology for more than five years.

BACKGROUND: Reverse transcriptase (RT) is present in all infectious retrovirus particles. Sensitive RT tests should thus detect all such particles. A family of ultrasensitive RT tests, product-enhanced reverse transcriptase (PERT) assays, have been designed. Accumulated results show that (i) a first version of the PERT assay that uses microtiter/ELISA technology detects RT of only 3-11 retroviral particles, (ii) very different human and animal lenti- and oncoviruses are detected very sensitively, (iii) HIV-1 is detected as sensitively as with polymerase chain reaction (PCR) for viral RNA, and (iv) prevalence of elevated particle-associated RT in plasma of unselected Swiss blood donors was 1.9%. OBJECTIVE: To investigate whether the RT activity detected in one unselected donor with a chronically elevated level of the liver enzyme alanine aminotransferase and two selected donors with chronically indeterminate or borderline-positive HIV-1 serology was due to human immunodeficiency virus (HIV) or human T-cell leukemia virus (HTLV) infection. STUDY DESIGN: Blood samples were tested by PCR for viral DNA and/or RNA of HIV-1, HIV-2, HTLV-1, HTLV-2, hepatitis B and C virus. Serological tests and virus cultures were also employed. RESULTS: Infection with any of the above agents could not be demonstrated. Virus cultivation in one case of borderline-positive HIV-1 Western blot for more than 5 years yielded a peak of RT production that was repeatedly transmissible to fresh cells. CONCLUSIONS: The findings suggest the presence of an infectious RT-positive agent, probably different from HIV-1/2 or HTLV-1/2, in a healthy individual with chronically borderline-positive HIV-1 serology. The PERT assay may detect retroviruses currently undetectable by other tests. The use of more stringent Western blot interpretation guidelines other than those of CDC or WHO is strongly recommended.

Journal Article↗

PubMed1996

Heat-mediated immune complex dissociation and enzyme-linked immunosorbent assay signal amplification render p24 antigen detection in plasma as sensitive as HIV-1 RNA detection by polymerase chain reaction.

OBJECTIVE: To compare heat denaturation and acidification as immune complex dissociation (ICD) methods in adult HIV-1 infection and to increase the sensitivity by a signal-amplification-boosted HIV-1 p24 antigen enzyme-linked immunosorbent assay (ELISA). DESIGN: Prospective and retrospective blinded study of paired serum and plasma samples from 245 patients (112 of class A, 66 of B, 67 of C of the Centers for Disease Control and Prevention 1993 classification). METHODS: Plasma and sera were prospectively tested for antigen by ELISA using native, acidified, or heat-denatured material. Retrospective tests included batch analysis of heat-denatured samples for antigen by the signal-amplification-boosted ELISA and for viral RNA. RESULTS: With serum, native antigen was reactive in 26.5%. Acidification increased the rate to 53.1% (P < or = 0.0001), but was inefficient at a CD4+ count > or = 500 x 10(6)/l. Heat denaturation further elevated the rate to 67.8% (P < or = 0.0007) and the use of plasma to 78.0% (P < or = 0.008). The boosted ELISA, performed with plasma samples diluted 1 :6, which eliminated the problem of heat-induced sample coagulation, was confirmed positive in 89.5% of serum and 97.8% of plasma samples. RNA was detected in 95.7%. CONCLUSION: Heat-mediated ICD combined with a signal-amplification-boosted ELISA detects HIV-1 expression as sensitively as a polymerase chain reaction kit for viral RNA, but at only a fraction of the cost. The procedure uses a 50 microliters plasma sample and should be fully automatable.

Adult↗

PubMed1994

Ultrasensitive retrovirus detection by a reverse transcriptase assay based on product enhancement.

Reverse transcriptase (RT) is an indispensable component of infectious retroviruses. We have developed an ultrasensitive RT test in which RNA of bacteriophage MS2 serves as the template for RT-mediated cDNA synthesis. A fragment of the cDNA is selectively amplified by polymerase chain reaction and the amplification product is analyzed by Southern blot hybridization or enzyme immunoassay. The procedure was 10(6) to 10(7) times more sensitive than a conventional RT test and detected as little as 10(-9) unit of murine leukemia virus RT, which corresponded to 2.1 x 10(2) molecules, a number present in 3-11 virions. As a screening assay for filterable particle-associated RT, it was positive with supernatants from cell cultures producing human immunodeficiency virus (HIV) type 1 or human T-cell leukemia virus (HTLV) type 1 or 2, but was negative with nonproducer cultures. It was positive with plasma samples from all tested individuals infected with HIV-1, HIV-2, or HTLV-1 and sera from cats infected with feline leukemia virus or feline immunodeficiency virus. Control samples from blood donors or uninfected cats were negative. Density banding experiments with culture supernatants showed that the RT activity was associated with virus particles. The assay should detect all replication-competent retroviruses or similar agents. It may be used as a screening assay for such agents, for quantitation of the viral load, drug susceptibility testing of RT, and control of virus inactivation in biological products.

Animals↗

PubMed1994

Sensitive detection and early prognostic significance of p24 antigen in heat-denatured plasma of human immunodeficiency virus type 1-infected infants. Swiss Neonatal HIV Study Group.

Immune complex formation causes underdetection of p24 antigen in human immunodeficiency virus (HIV) infection. Briefly boiling diluted plasma releases all complexed antigen, which can then be measured by some commercial assays. In a retrospective pediatric cohort study, the specificity of this procedure in 390 uninfected samples was 96.9% after initial testing and 100% after neutralization. Sensitivity among 125 postnatal infected samples was, at a detection of 2 pg/mL, 96.0% (97% neutralizable) compared with 47.7% for regular antigen (76% neutralizable), 96% for polymerase chain reaction, and 77% for viral culture. The high sensitivity and specificity of heat-denatured antigen was confirmed by prospectively testing 113 additional samples. Quantitative analysis of samples from infected infants showed low levels of p24 antigen in 29% of cord blood sera, a postnatal increase to levels that were during the first 6 months of life inversely associated with survival, and persistence of antigenemia thereafter independent of clinical status. Prevalence and antigen levels were significantly lower in mothers. The persistent antigenemia in children indicates that their immune systems cannot restrict HIV expression as efficiently as those of adults.

Age Factors↗

PubMed1994

A phase I/IIA clinical study with a chimeric mouse-human monoclonal antibody to the V3 loop of human immunodeficiency virus type 1 gp120.

A phase I/IIA clinical trial with the chimeric mouse-human monoclonal antibody CGP 47,439 to the principal neutralization determinant in the V3 region of human immunodeficiency virus type 1 (HIV-1) strain IIIB envelope protein gp120 is reported. The trial was an uncontrolled single-center, open-label, multidose tolerability, immunogenicity, and pharmacokinetic study in homosexual men with advanced HIV disease. Patient groups were formed on the basis of the reactivity of the antibody with the gp120 of their HIV-1 isolates. Intravenous infusions of 1, 10, and 25 mg of antibody were followed by seven escalated doses of 50, 100, and 200 mg, every 3 weeks. The antibody was well tolerated; no toxicity was observed. Some patients showed a transient but insignificant antibody response to the antibody with no apparent adverse reactions or accelerated elimination of it. Substantial serum levels of the antibody were maintained with a mean t1/2 beta of 8-16 days. A virus burden reduction was observed in some patients.

Acquired Immunodeficiency Syndrome↗

PubMed1994

IgG, IgM, and IgA response to HIV in infants born to HIV-1 infected mothers. Swiss Neonatal HIV Study Group.

Children born to HIV-1-positive mothers were prospectively tested for HIV-reactive IgG, IgM, and IgA by Western blot, in order to study the children's humoral immune response in the background of passively transferred maternal IgG. In infected infants, a response was first seen at 1-3 months for env-reactive IgM and IgA, as well as gag-reactive IgM and IgG. This was followed by production of IgG to env, IgA to pol and to gag p17 and p55 at 7-9 months, and IgG to pol at 10-12 months. IgG Western blot positivity by all interpretation guidelines in all infected infants was found by 10-12 months. Subsequently, only IgG to env and p24, and IgA to env were maintained in all, whereas IgG to pol and p17 disappeared again in a significant fraction. A considerable proportion of uninfected infants also produced gag-reactive antibodies: IgM at 1-3 months, followed by IgG, which persisted in 10-20% and were also found in children born to uninfected mothers. These antibodies were, however, present at lower titers than in infected infants and were apparently produced in response to agent(s) different from HIV. Maternal antibodies to env disappeared significantly faster in infected than uninfected infants. Traces of HIV-reactive IgG were present for up to 21 months in children who subsequently seroreverted completely.

Age Factors↗

PubMed1993

Sensitive and quantitative detection of PCR-amplified HIV-1 DNA products by an enzyme linked immunoassay following solution hybridization with two differently labelled oligonucleotide probes.

We have developed and evaluated an ELISA-based detection method for PCR-amplified HIV-1 DNA. The assay uses two oligonucleotide probes which are end-labelled at the 5'-end with biotin or digoxigenin, respectively. Upon solution hybridization of these probes which react with the same strand of amplified DNA product, the formed hybrids are bound to avidin-coated wells of a microtitre plate and detected by horseradish peroxidase-labelled antibodies directed against digoxigen and 3, 3', 5, 5'-tetramethylbenzidine as substrate. Factors critical for a high signal-to-noise ratio were the use of serum as blocking agent, the amount of biotin-labelled and digoxigenin-labelled oligonucleotide probes present in a reaction and the inactivation of Taq DNA polymerase. The method has a detection limit of 1-3 pg of amplified DNA and is quantitative in a range extending from 1 pg to at least 200 pg. In the background of 1 microgram of total DNA, one single-stranded copy of HIV-1 DNA can be detected after 35 cycles of amplification. A comparison of the ELISA-based detection method with primer extension analysis, a method previously shown to reach a similar detection limit, demonstrated complete agreement of the results of 118 amplified DNAs. The method proved simple, requires only about 3 h, and could easily be adapted to the detection of other amplified target DNAs.

Base Sequence↗

PubMed1993

Pharmacokinetics of an HIV-1 gp120-specific chimeric antibody in patients with HIV-1 disease.

The pharmacokinetics of mouse V/human C (gamma 1, kappa) chimeric monoclonal antibody CGP 47 439 specific for the principal neutralizing determinant of human immunodeficiency virus type 1 (HIV-1) was studied in patients with stage IV HIV-1 disease in an open-labeled phase I/IIA trial. Twelve male patients were enrolled and nine completed the study. Patients were divided into three groups according to the extent of CGP 47 439 to bind to gp120 from their viral isolates: undetectable for group 1, modestly reactive for group 2, and strongly reactive for group 3. A first dose of 1, 10, or 25 mg was administered by intravenous infusion to group 1, group 2 and group 3 patients, respectively. The patients then received seven doses of 50, 100, or 200 mg, respectively, every three weeks. CGP 47 439 serum concentrations were determined by an ELISA using monoclonal antibody AB19-4 specific for the idiotope of CGP 47 439. Half an hour after infusion only 25.5-36.1% of the administered antibody was found in the serum, reflecting its rapid distribution in the extravascular space and possibly binding to gp120 antigen in some of the patients. The terminal elimination half-life (T1/2) was 16.2 days in group 1 patients, 9.7 days in group 2 and in group 3 patients 7.5 days and 9.1 days. An antibody response to CGP 47 439 was not a factor in determining elimination rates, because only very low and transient responses were found in three patients. These results suggest that the reactivity of CGP 47 439 with HIV-1 gp120 contributed to its elimination in HIV-1 infected patients.

Adult↗

PubMed1993

Primer extension analysis provides a sensitive tool for the identification of PCR-amplified DNA from HIV-1.

Primer extension analysis was evaluated as a means to identify PCR-amplified DNA from HIV-1. Solution hybridization with radioactive labeled oligonucleotides followed by an extension reaction with Klenow fragment of Escherichia coli DNA polymerase I and subsequent separation on denaturing polyacrylamide gels reveals single stranded DNA products of the predicted size. The specificity of the reaction is further demonstrated by specific endonuclease digestion. The analysis is more sensitive than Southern blotting and about equally sensitive as Slot blot analysis when double-stranded DNA probes are used for hybridization. With end-labeled oligonucleotide probes, primer extension analysis proved an order of magnitude more sensitive than membrane hybridization. The analysis also allows quantitation of amplified DNA from 1 pg to about 1 ng of DNA product. Under the conditions described for amplification, primer extension analysis is capable of detecting a single HIV-1 plasmid DNA molecule in the presence of 1 microgram of total DNA. 3'-end mismatching of the oligonucleotide probe does not result in a significantly altered detection limit. Primer extension analysis can also be carried out with at least three different PCR-amplified DNAs in the same reaction tube.

Base Sequence↗

PubMed1993

Quantitative and sensitive detection of immune-complexed and free HIV antigen after boiling of serum.

5-min boiling of 3-fold diluted serum leads to the quantitative release of immune-complexed HIV antigen (Ag) that can be readily measured. The method is capable of detecting as little as 4 to 5 pg/ml Ag p24 depending on the Ag test used. Of a panel of 50 anti-HIV-positive sera, 30 (60%) were Ag-positive after heat denaturation, 17 (34%; P < or = 0.01) after acidification, and 13 (26%; P < or = 0.001) when tested undenatured. The superiority of heat denaturation was greatest in sera with high-titered antibodies to p24. Occurrence and mean concentrations of heat-denatured Ag were reversely correlated with p24-specific antibody. It is suggested that heat-denatured proteins be used for raising antibodies that will serve as capture or trace antibodies in Ag assays. This simple change in the manufacturing of test reagents in combination with a short boil before testing may solve the problem of incomplete Ag detection due to immune complex formation.

Antigen-Antibody Complex↗

PubMed1993

PCR identification of HIV-1 DNA sequences in brain tissue of patients with AIDS encephalopathy.

We analyzed brain tissue from 39 patients for the presence of proviral HIV-1 sequences, using the polymerase chain reaction (PCR) for the amplification of segments of the viral LTR and gag genes. A novel primer extension procedure allowed the detection of a single HIV-1 copy in 1 micrograms DNA. We detected proviral HIV-1 DNA in 16 of 25 brain samples from AIDS patients. Semiquantitative evaluation of the amplified DNAs indicated considerable variation in viral load. Highest levels of proviral DNA were present in brain samples from six patients with clinical evidence of HIV-associated cognitive/motor complex and the histopathologic correlate of HIV leukoencephalopathy or HIV encephalitis. An additional 11 brain samples contained smaller amounts of proviral DNA. In these patients, clinical data were inconclusive regarding the diagnosis of HIV-1 encephalopathy and histopathologically there was no evidence of HIV-1-induced tissue lesions. In nine of 25 seropositive patients with AIDS (36%), brain samples scored negative or did not contain an unequivocal signal indicating the presence of proviral DNA. HIV-1 sequences were not detected in any of 14 control brain samples from HIV-1 seronegative patients. Our data indicate that HIV-1 is present in the central nervous system of the majority (two thirds) of AIDS patients and that the highest levels of proviral DNA in brain tissue are associated with HIV encephalopathy.

AIDS Dementia Complex↗