Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UMLS:C0021051 (immunodeficiency)
 71,517 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The ROD strain of the human immunodeficiency virus type 2 (HIV-2) was used to produce monoclonal antibodies. Virus grown in CEM cells was partially purified by ultracentrifugation and solubilized in a buffer containing Triton X-100. BALB/c mice were inoculated intraperitoneally with 50 micrograms of solubilized virus preparations mixed 1:1 with complete Freund's adjuvant. Animals were boosted on day 28 and sacrificed on day 31. Spleen cells from the immunized animals were fused with SP20/Ag 14 myeloma cells and cultured in HAT medium. Following selection of the hybrids of interest by an HIV-2 ELISA procedure, hybridomas were cloned twice by limiting dilution. Six clones were found to produce antibodies that reacted with HIV-2 antigens as judged by ELISA. These antibodies were concentrated by ammonium sulfate precipitation, and analyzed by the Western blot procedure. Monoclonal antibodies specifically reactive to an HIV protein of 68 KD were obtained. These antibodies did not react with an HIV-2 band of 55 KD. These data showed that the monoclonal antibodies recognized the carboxy terminal region (the RNAse H domain) of the HIV-2 retrotranscriptase enzyme.
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PMID:Production of monoclonal antibodies to human immunodeficiency virus type-2. 128 63

Fresh frozen plasma (FFP) is prepared in blood banks world-wide as a by-product of red blood cell concentrate preparation. Appropriate clinical use is for coagulation factor disorders where appropriate concentrates are unavailable and when multiple coagulation factor deficits occur such as in surgery. Viral safety depends on donor selection and screening; thus, there continues to be a small but defined risk of viral transmission comparable with that exhibited by whole blood. We have prepared a virus sterilized FFP (S/D-FFP) by treatment of FFP with 1% tri(n-butyl)phosphate (TNBP) and 1% Triton X-100 at 30 degrees C for 4 hours. Added reagents are removed by extraction with soybean oil and chromatography on insolubilized C18 resin. Treatment results in the rapid and complete inactivation of greater than or equal to 10(7.5) infectious doses (ID50) of vesicular stomatitis virus (VSV) and greater than or equal to 10(6.9) ID50 of sindbis virus (used as marker viruses), greater than or equal to 10(6.2) ID50 of human immunodeficiency virus (HIV), greater than or equal to 10(6) chimp infectious doses (CID50) of hepatitis B virus (HBV), and greater than or equal to 10(5) CID50 of hepatitis C virus (HCV). Immunization of rabbits with S/D-FFP and subsequent adsorption of elicited antibodies with untreated FFP confirmed the absence of neoimmungen formation. Coagulation factor content was comparable with that found in FFP. Based on these laboratory and animal studies, together with the extensive history of the successful use of S/D-treated coagulation factor concentrates, we conclude that replacement of FFP with S/D-FFP, prepared in a manufacturing facility, will result in improved virus safety and product uniformity with no loss of efficacy.
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PMID:Solvent/detergent-treated plasma: a virus-inactivated substitute for fresh frozen plasma. 131 64

Deletions were constructed within a functional human immunodeficiency virus type 1 (HIV-1) proviral clone in order to assess the role of the envelope protein in virus particle formation. A graded exonuclease deletion technique was used to produce 12 clones with deletions of 175-308 nucleotides in the first conserved domain of envelope. This included 9 clones with frameshift deletions and 3 clones with in-frame deletions. Isogenic pairs of env deletion clones were produced with or without an additional deletion in the vif and vpr genes. Upon transfection, all clones produced virus particles, as determined by p24 antigen, reverse transcriptase, and sucrose gradient assays with conditioned media. Virus particles produced from clones with deletions in env or vif and vpr, or both regions, banded on sucrose gradients with a mobility similar to that of virus produced by the parental clone. The p24 gag capsid protein in the particles was resistant to trypsin, but the particles were disrupted by treatment with Triton X-100, suggesting the presence of a surrounding lipid bilayer. Furthermore, electron microscopic studies revealed both mature and immature virus particles derived from COS cells transfected with the env deletion clones. Cocultivation experiments with lymphoid cells and cells transfected with each of the env deletion clones demonstrated that the virus particles were noninfectious.
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PMID:Formation of noninfectious HIV-1 virus particles lacking a full-length envelope protein. 182 17

An 80-kilodalton glycoprotein (gp80) was produced in human immunodeficiency virus type 2 (HIV-2)-infected cells along with three envelope glycoproteins that we have recently reported: the extracellular glycoprotein (gp125), the envelope glycoprotein precursor (gp140), and the transient dimeric form of the precursor (gp300). gp125 and gp80 were detectable after the synthesis of gp140 and the formation of gp300. Using a specific monoclonal antibody, we showed here that gp80 is a dimeric form of the transmembrane glycoprotein gp36 of HIV-2. Dimerization of the envelope glycoprotein precursor and dimeric forms of the transmembrane glycoproteins were also observed in cells infected with simian immunodeficiency virus (SIV-mac), a virus closely related to HIV-2. Under routine conditions of our experiments (i.e., extraction by 1% Triton X-100 before polyacrylamide gel electrophoresis in sodium dodecyl sulfate [SDS]), monomeric forms of the transmembrane glycoprotein of HIV-2 and SIV-mac were only seldomly observed. Dimeric forms of the envelope precursors and the transmembrane glycoproteins are probably stabilized by extraction in the nonionic detergent Triton X-100 since such dimeric forms resist dissociation during subsequent electrophoresis in the presence of the ionic detergent SDS. However, the dissociation of these dimeric forms might occur when samples are prepared by extraction directly in 1% SDS or by incubation of the purified dimers at acidic pH. Dimerization of the envelope precursor might be required for its processing to give the mature envelope proteins, whereas the transmembrane dimer might be essential for optimal structure of the virion and thus its infectivity.
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PMID:Transmembrane envelope glycoproteins of human immunodeficiency virus type 2 and simian immunodeficiency virus SIV-mac exist as homodimers. 229 88

A simple, sensitive, and specific polymerase chain reaction (PCR) protocol for the detection of human immunodeficiency virus type 1 (HIV-1) is described. We have improved all three PCR steps: sample preparation, DNA amplification, and detection of the amplified product. Some of the improvements have been described previously, but they have never been combined into a complete PCR protocol. Peripheral blood mononuclear cells were lysed directly in a buffer containing sodium dodecyl sulfate, Triton X-100, and proteinase K. This crude cell lysate was amplified in a two-step PCR, first with outer primers and then with inner primers nested within the first primers. The PCR product was visualized by agarose gel electrophoresis and ethidium bromide staining. Thus, we avoided conventional DNA extraction as well as hybridization for the detection of the PCR product. The samples were analyzed with four sets of nested primers (JA4 through JA7, JA9 through JA12, JA13 through JA16, and JA17 through JA20) designed to amplify HIV-1 gag, env gp120, env gp41, and pol sequences, respectively. We were able to amplify HIV-1 sequences in all samples from 90 HIV-1-seropositive individuals with mostly mild symptoms. Of these individuals, 24 were negative in HIV-1 isolation and 9 were selected because they were infected by African and Haitian HIV-1 strains. Eighty-five (94%) individuals were positive with at least three of four primer sets. Samples from 26 healthy blood donors, as well as cells infected in vitro with human immunodeficiency virus type 2 and human T-cell leukemia virus type I, were negative in PCR, thus demonstrating the specificity of the amplification.
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PMID:Simple, sensitive, and specific detection of human immunodeficiency virus type 1 in clinical specimens by polymerase chain reaction with nested primers. 238 Mar 80

The treatment of plasma with organic solvent/detergent mixtures at the time of plasma collection or pooling could reduce the exposure of technical staff to infectious viruses and enhance the viral safety of the final product. Treatment of plasma for 4 hours with 2-percent tri(n-butyl)phosphate (TNBP) at 37 degrees C, with 1-percent TNBP and 1-percent polyoxyethylensorbitan monooleate (Tween 80) at 30 degrees C, or with 1-percent TNBP and 1-percent polyoxyethylene ethers, (Triton X-45) at 30 degrees C resulted in the rapid and complete inactivation of greater than or equal to 10(4) tissue culture-infectious doses (TCID50) of vesicular stomatitis and Sindbis viruses, which are used as surrogates. Treatment of plasma with TNBP and TNBP and Tween-80 was shown to inactivate greater than or equal to 10(4) TCID50 of human immunodeficiency virus. TNBP treatment of plasma contaminated with 10(6) chimpanzee-infectious doses (CID50) of hepatitis B virus and 10(5) CID50 of non-A,non-B hepatitis virus prevented the transmission of hepatitis to chimpanzees. Immediately after treatment of plasma with 2-percent TNBP, the recovery of factors VIII, IX, and V and antithrombin III was 80, 90, 40, and 100 percent, respectively. Recovery of all factors was greater than or equal to 90 percent after treatment with TNBP and detergent mixtures. Treated plasma was fractionated by standard techniques into antihemophilic factor and prothrombin complex concentrates, immune globulin, and albumin. Prior treatment with TNBP or TNBP and detergent did not affect the separations of desired proteins. Therefore, it appears possible to inactivate viruses in plasma before the execution of standard fractionation procedures.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:The use of tri(n-butyl)phosphate detergent mixtures to inactivate hepatitis viruses and human immunodeficiency virus in plasma and plasma's subsequent fractionation. 175 94

Flow cytometric detection of human immunodeficiency virus (HIV)-infected lymphoid cells at low frequencies is described. Infected cells from human T lymphoid cell lines H9 and A3.01 were detected at frequencies as low as 10(-4) following indirect immunofluorescence labeling. For labeling, cells were treated with an HIV-inactivating, permeabilizing fixative followed by binding of a monoclonal antibody specific for the HIV major core protein p24, and then by binding of fluorescein isothiocyanate-conjugated F(ab')2 fragments of goat anti-mouse immunoglobulin antibody. We compared two fixation procedures, one using a mixture of methanol and acetone, the other a three-step fixation using methanol, paraformaldehyde and Triton X-100. The latter fixation protocol was found to be superior in its ability to resolve mixtures of infected and uninfected cells. The method allowed determination of the percentage of the cell population that was infected and the relative amount of p24 antigen per cell. At analysis rates of several thousand cells/s, detection of HIV-infected cells as rare events was possible. Excellent agreement was obtained between flow cytometric evaluation and reverse transcriptase (RT) assay of infected H9 cells cocultured with uninfected H9 cells in various proportions for 7 days. In time course of infection experiments, cultures infected by small numbers of viral particles were positive by flow cytometry up to 3 days earlier than by RT assay.
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PMID:Detection of human immunodeficiency virus-infected lymphoid cells at low frequency by flow cytometry. 244 28

CD4 is an integral membrane glycoprotein that acts as the cellular receptor for human immunodeficiency virus (HIV). A cDNA encoding full-length CD4 was inserted into the genome of Autographa californica nuclear polyhedrosis virus under transcriptional regulation of the viral polyhedrin gene promoter. The recombinant virus was used to infect insect cells, which resulted in the abundant expression of CD4 as evaluated by flow cytometry and immunoblot analysis. Recombinant CD4 expressed on the surface of infected insect cells was immunologically indistinguishable from human CD4 when using 11 different anti-CD4 monoclonal antibodies. The extraction of infected cells by phase-transition separation with Triton X-114 followed by immunoaffinity chromatography yielded a single protein detected by NaDodSO4/PAGE using silver staining. N-terminal sequence analysis of the purified recombinant protein showed that CD4 produced in Sf9 cells is efficiently cleaved from the precursor protein. Immunoblot analysis under nondenaturing conditions showed that the purified protein reacted with the anti-CD4 monoclonal antibody Leu-3a. The potential use of the recombinant membrane-associated CD4 in anti-HIV therapy is discussed.
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PMID:Cell-surface expression and purification of human CD4 produced in baculovirus-infected insect cells. 268 21

Simple polyesters derived from poly(ethylene glycol)s and alpha, omega-dicarboxylic acids exhibit a broad range of activity in disrupting phospholipid membranes. This activity has been analyzed by measuring the release of liposome-encapsulated 5(6)-carboxy-fluorescein (CF). Comparison with an analogous monomeric surfactant, and with Triton X-100, demonstrates that macromolecular activity is a sensitive function of the size of the hydrophobic and hydrophilic segments within each repeat unit, and that high disrupting power is possible. In vitro studies with the human immunodeficiency virus type-1 have revealed that those polyesters which exhibit the highest membrane disrupting power also provide significant protection for human CD4+ lymphocytes against HIV-1. The potential for adjusting and utilizing these "supramolecular surfactants" in medicine is briefly discussed.
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PMID:Supramolecular surfactants: amphiphilic polymers designed to disrupt lipid membranes. 293 May 30

Since the human immunodeficiency virus (HIV) may be transmitted accidentally to laboratory personnel analyzing patient sera, the efficiency of a non-ionic detergent, Triton X-100, in inactivation of HIV in human serum as a safety measure was studied. Semliki Forest virus, an enveloped toga virus, was used as a model virus to create optimal treatment conditions. In the presence of 50% serum, complete inactivation (i.e. no residual virus detected, greater than 7 log reduction of virus titre) was achieved by incubation with 0.2% Triton X-100 for 1 h at 37 degrees C. Under these conditions HIV was also completely inactivated (i.e. no residual infectious virus detected, greater than or equal to 5 log reduction of virus titre). Both treated and untreated serum specimens were also tested with several enzyme immunoassays used in virological laboratories to determine whether the inactivation treatment interfered with the assays. The treated specimens, further diluted as recommended for each assay, were subjected to 15 enzyme immunoassays for microbial antibodies and antigens (HIV IgG, hepatitis A IgG and IgM, hepatitis B s, c, and e antigens and antibodies, cytomegalovirus IgG, mumps virus IgG, poliovirus IgG, rubellavirus IgM, toxoplasma IgG, and chlamydia IgG). Clearly decreased sensitivity was found only with two hepatitis B tests (e antigen and antibody to the surface antigen). It is concluded that safe inactivation of HIV in serum is achieved by 0.2% Triton X-100, but the treatment may decrease the sensitivity of some tests in which low specimen dilution is used.
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PMID:Inactivation of human immunodeficiency virus in serum specimens as a safety measure for diagnostic immunoassays. 314 Nov 60


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