ABSTRACT
OBJECTIVES: To develop a tool for then easy and user-friendly management of peptide microarray experiments and for the use of the results of these experiments for the study the immune response against HIV virus infection in clinical samples. METHODS: Applying bioinformatics and statistics for the analysis of data coming from microarray experiments as well as implementing a MIAME (Minimum Information About a Microarray Experiment) compliant database for managing and annotating experiments, results and samples. RESULTS: We present a new tool for managing not only nucleic acid microarray experiments but also protein microarray experiments. From the analysis of experimental data, we can detect different profiles in the reactivity of the sera with different genotypes. CONCLUSIONS: We have developed a new tool for managing microarray data including clinical annotations for the samples as well as the capability of annotating other microarray formats different to those based on nucleic acids. The use of peptide microarrays and bioinformatics analysis opens a new scope for the characterization of the immune response, and analyzing and identifying the humoral response of viruses with different genotypes.
Subject(s)
HIV Seropositivity/genetics , Oligonucleotide Array Sequence Analysis , Peptides/genetics , Viral Proteins/genetics , Computational Biology , HIV Antibodies/biosynthesis , HIV Seropositivity/immunology , Humans , Oligonucleotide Array Sequence Analysis/statistics & numerical data , Protein Array Analysis , Serotyping , Spain , Statistics as Topic , Viral Proteins/chemistryABSTRACT
We have evaluated the prevalence of HIV-1 non-B subtypes in Spain by means of an enzyme immunoassay (EIA) for discrimination between B and non-B subtypes. Samples were obtained from newly diagnosed patients attended at internal medicine outpatient clinics between October 1997 and October 1998. Discrimination between HIV-1 B and non-B subtypes was carried out by means of the EIA, with V3 synthetic peptides specific to the different subtypes. Non-B-serotyped samples were genetically analyzed in the gp41 region from the original sera. During the study period, 909 samples were collected from 21 medical units located in various Spanish geographical regions. Serotyping was possible in 885 cases, of which 791 were assigned as B serotype (89.38%), 70 showed no reactivity to any of the peptides (7.91%), and the remaining samples displayed other reaction patterns (2.72%). Of the 94 non-B-assigned samples, 65 were genetically characterized in the gp41 region of the env gene: 55 were B subtype, 5 were A subtype (4 clustered with CRF02AG reference strains), 3 were C subtype, and 2 were G subtype. The prevalence rate for non-B subtypes in Spain was established at 1.13% (95% CI, 0.59-2.21). Although the B subtype is predominant in the Spanish population, other subtypes have been detected.
Subject(s)
Enzyme-Linked Immunosorbent Assay/methods , HIV Infections/virology , HIV-1/classification , Gene Products, env/analysis , HIV Envelope Protein gp41 , HIV Infections/epidemiology , Humans , Peptide Fragments , Prevalence , Serotyping , Spain/epidemiologyABSTRACT
A variant of the murine CD4+ T helper cell clone D10.G4.1 (D10) has been isolated and cloned. This line, which we have named "syngeneic-reactive D10", or SR.D10, maintains the I-Ak-restricted specificity for Conalbumin and the allogeneic specificities characteristic of D10 cells. However, it is hyperreactive to TCR-dependent and-independent stimuli, indicating a lower activation threshold than the original D10.G4.1 clone. The hyperreactivity of SR.D10 runs in parallel with the acquisition of a reactive phenotype against syngeneic antigen presenting cells (APCs). As in antigen activation, reactivity to syngeneic APCs can be inhibited by anti-TCR, anti-CD4, or anti-class II monoclonal antibodies. The role of CD4 in this phenomenon is highlighted as "syngeneic reactivity" disappears in CD4- mutants of SR.D10 and is recovered in CD4 transfectants. The expression of several cell surface molecules involved in T cell activation show qualitative and/or quantitative differences between SR.D10 and the original D10. No significant differences in quantity and activity of p56lck and p59fyn were detected between the hyperreactive and the original clone. Our results suggest that high sensitivity to activation, concomitant with expression of CD4, might allow the acquisition of an autoreactive phenotype and confirm the important contribution of coreceptors to determine the activation threshold of the cells. The characteristics of SR.D10 and the possibility of growing them in the presence of interleukins make this cell line a experimental model of great interest for analyzing activation mechanisms in T cells.
Subject(s)
Antigen-Presenting Cells/immunology , CD4-Positive T-Lymphocytes/immunology , Lymphocyte Activation , Animals , Antigens, Differentiation, T-Lymphocyte/metabolism , CD4 Antigens/physiology , Cell Line , Cell Separation , Histocompatibility Antigens Class II/physiology , Immunophenotyping , Mice , Mice, Inbred Strains , Receptors, Antigen, T-Cell/physiologyABSTRACT
Cross-reactions of 17 members of the family Legionellaceae were studied by four different serological techniques: immunofluorescence (IF), slide agglutination (SA), microagglutination (MA) and immunodiffusion (ID), using antigens and rabbit antisera prepared in our laboratory. Results obtained corresponded closely with those described by other authors, especially for IF and SA. The 17 antigens were further tested by IF with a panel of sera previously diagnosed as positive for legionella. A high number of positive reactions with several of the antigens tested were found, half of them being positive for Legionella pneumophila serogroup 1, usually in combination with other serogroups or species. The remaining sera presented a great variety of patterns combining different antigens.
