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51.
Dr. David Lo Christina R. Reilly Linda C. Burkly Jenefer DeKoning Terri M. Laufer Laurie H. Glimcher 《Immunologic research》1997,16(1):3-14
Ten years ago, we proposed a model for thymus function in which thymic epithelial cells are primarily responsible for imprinting
major histocompatibility complex (MHC)-restricted specificity, and bone marrow-derived macrophages or dendritic cells are
responsible for the induction of self-tolerance. Since then, transgenic and knockout models have allowed for a dissection
of thymic stromal components in vivo, leading to a new understanding of their specialized functions. We have determined that
with regard to class II-restricted CD4 T-cell development, two distinct subsets of thymic epithelium help shape the repertoire:
Cortical epithelium appears solely responsible for positive selection, whereas a fucose-bearing subset of medullary epithelium
is specialized for negative selection. This absolute separation of positive and negative selection into two distinct spatial
and temporal compartments leads to a much simpler view of the process of repertoire selection. Finally, a novel view of the
function of the thymic medulla is discussed. 相似文献
52.
Summary. Nested polymerase chain reaction (PCR) amplifying the morphological transforming region II (mtrII) of cytomegalovirus (CMV)
has been shown to be useful in the detection of CMV DNA in bone marrow transplant (BMT) recipients. However, there has never
been any report on mutation hot spots and subtypes of this open reading frame. Using primers derived from sequences upstream
and downstream of mtrII (ORF 79), CMV DNA from peripheral blood leukocytes (PBL) and conventional CMV culture of 16 BMT recipients
were amplified by PCR, cloned into pUC118, and sequenced. The amino acid sequences were predicted using the standard triplet
code. The DNA sequences obtained from direct amplification of CMV in PBL obtained from the 16 patients were 100% identical
to the corresponding ones obtained by amplification of CMV DNA extracted from conventional CMV culture. Within mtrII (ORF
79), hot spot single base mutations were observed at positions +40 (G→A), +123 (A→G), +213 (T→C), and +219 (T→C). However,
because of third base degeneracy, only amino acid 14 was changed from valine to isoleucine in the predicted protein of 13
patients. This corresponded to the hot spot mutation at position +40 (GTC→ATC), while the rest were silent mutations. An insertion
of 3 bases (ACG) was observed in the CMV DNA of 10 patients at positions +91 to +93, leading to a threonine insertion at amino
acid 31 in these patients. For patient no. 147 there was a 65 bp deletion in the CMV DNA amplified later in the course of
BMT as compared with that early in the course. This gave rise to a frame shift mutation and a change of more than 70% in the
predicted amino acid sequence of the protein.
Accepted October 14, 1998 Received May 20, 1998 相似文献
53.
R C Porter P Lo D E Low A E Simor A McGeer S Scriver T C Moore C Goldman M Skulnick 《Journal of clinical microbiology》1993,31(10):2794-2795
The BACTEC PLUS 26 (NR26) (Becton Dickinson, Towson, Md.) high-volume blood culture bottle replaced the less expensive smaller-volume NR6A bottle in our hospital. An audit carried out several months after their introduction revealed that only 17.5% of the NR26 bottles received the required blood volume. Several audits and educational programs were required in order to achieve a compliance rate of > 60%. 相似文献
54.
AIM: Development of a specific polymerase chain reaction (PCR) assay for detection of the pre-core, stop codon, mutant of hepatitis B virus (HBV). METHODS: PCR primers, specific at the 3'-end for nucleotide 1896 of either the pre-core, stop codon, mutant or wild type HBV, were synthesised using published sequence data. Positive control templates for both types of virus were synthesised by the PCR, incorporating sequences specific for each virus type at the appropriate position. These templates were used to optimise the specificity of the procedure. Formalin fixed, paraffin wax embedded human tissue from acute or fulminant HBV hepatitis from Hong Kong or Oxford was then investigated for presence of mutant or wild type virus. The HBV DNA was amplified from this tissue using a two step procedure, with an initial amplification phase followed by a second diagnostic phase on optimally diluted target DNA. RESULTS: Specific detection of mutant or wild type HBV was achieved. An important factor in determining specificity was the temperature of annealing, 70 degrees C proving to be highly specific. To overcome the inherent variation of target copy number in clinical samples and to provide an intrinsic positive control, it was important to generate and standardise the amount of target HBV used for the specific PCR. Two cases of fulminant hepatitis and four cases of acute hepatitis from Hong Kong, and one case of fulminant hepatitis from Oxford, contained only wild type HBV, with no evidence of a mutant virus. CONCLUSION: This method can be applied to FFPE tissues. It is rapid, non-radioactive, and specific for the stop codon mutation at nucleotide 1896 of HBV. Preliminary investigation of a small number of cases of fulminant hepatitis from Oxford and Hong Kong showed only wild type virus. The result differs from results published from Japan and Israel. 相似文献
55.
56.
Rapid Cytomegalovirus pp65 Antigenemia Assay by Direct Erythrocyte Lysis and Immunofluorescence Staining 总被引:3,自引:3,他引:3 下载免费PDF全文
Stephen K. N. Ho Chi-Yuen Lo Ignatius K. P. Cheng Tak-Mao Chan 《Journal of clinical microbiology》1998,36(3):638-640
A rapid cytomegalovirus (CMV) pp65 antigenemia assay with direct erythrocyte lysis (DL) with 0.8% NH4Cl, followed by indirect immunofluorescence staining (IF), was evaluated with 82 blood samples from renal transplant recipients, and the results were compared to those of the conventional antigenemia assay with dextran sedimentation and two-cycle alkaline phosphatase, anti-alkaline phosphatase staining (DS-APAAP). The DL-IF modification gave a higher leukocyte yield compared to DS-APAAP (75.4 versus 54.9%; P < 0.05), with similar leukocyte viability rates of >95%. The DL-IF methodology involved fewer technical steps, and the assay time was shortened from 5 h to less than 3 h. Nineteen of the 82 samples concordantly tested positive for pp65 antigenemia by both assays, and the readings showed a good correlation (r = 0.996; P < 0.01). No discordant results were observed. We conclude that the CMV pp65 antigenemia assay by this novel DL-IF modification is technically simpler, cheaper, and less time-consuming but yields results comparable to those of the conventional DS-APAAP assay. The shortened assay time and increased capacity to handle more samples confer distinct advantages in the rapid diagnosis and prompt treatment of CMV disease in immunosuppressed patients. 相似文献
57.
Saulot V Vittecoq O Salle V Drouot L Legoedec J Le Loët X Godin M Ducroix JP Ménard JF Tron F Gilbert D 《Journal of autoimmunity》2002,19(1-2):55-61
To identify new autoantibody populations in patients with rheumatic diseases, a cDNA expression library was immunoscreened with a rheumatoid arthritis (RA) patient's serum which contains autoantibodies binding to uncharacterized polypeptides by Western-blotting. One clone encoding the amino-terminal region (Nt) [domain L and half of domain I] of human calpastatin was selected. Different fragments of the selected cDNA were prepared and the corresponding recombinant polypeptides were produced by in vitro translation and analysed by Western blotting. Most RA sera bound to recombinant amino-terminal region and domain I but not to domain L. This prompted us to use a recombinant polypeptide corresponding to the domain I of calpastatin as the antigen in a solid-phase ELISA to test sera from patients with various systemic rheumatic diseases and healthy controls.Anti-calpastatin domain I antibodies (ACAST-DI Ab), were detected by ELISA in RA, systemic lupus erythematosus (SLE), Sj?gren's syndrome and control sera at respective frequencies of 10, 9, 0 and 1%. These Ab did not have prognostic value in early RA; high levels were significantly associated with vasculitis in SLE. Antibodies reacting with the calpastatin amino-terminal region are produced during systemic rheumatic diseases and are predominantly directed against domain I. High levels of these Ab may constitute a marker of vasculitis in SLE. 相似文献
58.
The first clinical isolate of Legionella parisiensis, from a liver transplant patient with pneumonia. 总被引:1,自引:0,他引:1 下载免费PDF全文
F Lo Presti S Riffard F Vandenesch M Reyrolle E Ronco P Ichai J Etienne 《Journal of clinical microbiology》1997,35(7):1706-1709
A bluish white autofluorescent strain of Legionella was isolated from the tracheal aspirate of a female liver transplant patient who developed hospital-acquired pneumonia. This strain had biochemical characteristics compatible with those of L. cherrii, L. anisa, and L. parisiensis and could not be differentiated from L. bozemanii and L. parisiensis by the direct fluorescent-antibody assay. Phylogenetic analysis of partial 16S rRNA gene sequences of this strain (ATCC 700174) revealed the closest homology to the species L. parisiensis (99.5%). An L. parisiensis species-specific profile was also identified by a random amplified polymorphic DNA technique. This is the first report of L. parisiensis isolation from humans. 相似文献
59.
Detection and Identification of Actinobacillus pleuropneumoniae Serotype 5 by Multiplex PCR 总被引:9,自引:0,他引:9 下载免费PDF全文
Terry M. Lo Christine K. Ward Thomas J. Inzana 《Journal of clinical microbiology》1998,36(6):1704-1710
Serotyping of Actinobacillus pleuropneumoniae is based on detection of the serotype-specific capsular antigen. However, not all isolates can be serotyped, and some may cross-react with multiple serotyping reagents. To improve sensitivity and specificity of serotyping and for early detection, a multiplex PCR assay was developed for detection of A. pleuropneumoniae and identification of serotype 5 isolates. DNA sequences specific to the conserved export and serotype-specific biosynthesis regions of the capsular polysaccharide of A. pleuropneumoniae serotype 5 were used as primers to amplify 0.7- and 1.1-kb DNA fragments, respectively. The 0.7-kb fragment was amplified from all strains of A. pleuropneumoniae tested with the exception of serotype 4. The 0.7-kb fragment was not amplified from any heterologous species that are also common pathogens or commensals of swine. In contrast, the 1.1-kb fragment was amplified from all serotype 5 strains only. The assay was capable of amplifying DNA from less than 102 CFU. The A. pleuropneumoniae serotype 5 capsular DNA products were readily amplified from lung tissues obtained from infected swine, although the 1.1-kb product was not amplified from some tissues stored frozen for 6 years. The multiplex PCR assay enabled us to detect A. pleuropneumoniae rapidly and to distinguish serotype 5 strains from other serotypes. The use of primers specific to the biosynthesis regions of other A. pleuropneumoniae serotypes would expand the diagnostic and epidemiologic capabilities of this assay. 相似文献
60.
Expression of sialyl-Lewis X, an E-selectin ligand, in inflammation, immune processes, and lymphoid tissues. 总被引:1,自引:0,他引:1 下载免费PDF全文
J. M. Munro S. K. Lo C. Corless M. J. Robertson N. C. Lee R. L. Barnhill D. S. Weinberg M. P. Bevilacqua 《The American journal of pathology》1992,141(6):1397-1408
The carbohydrate structure sialyl-Lewis X (SLex) can function as a ligand for E-selectin, formerly known as endothelial leukocyte adhesion molecule-1 (ELAM-1). This study was performed to analyze the expression of SLex by leukocytes and other cell types in the context of inflammatory and immune processes. Human peripheral blood cells were examined by flow cytometry using monoclonal antibody CSLEX1 directed against SLex. Cell surface SLex was found in abundance on nearly all isolated polymorphonuclear leukocytes (PMN) and monocytes, and at low levels on a substantial portion (up to 40%) of natural killer cells. This moiety was expressed also on approximately 10% of peripheral blood T cells. Immunohistochemistry was performed on various human tissues involved in inflammatory or immune processes and on secondary lymphoid tissues. In acute appendicitis, endothelial cells of postcapillary venules expressed E-selectin, and most PMN, both within vessels and extravasated, expressed SLex. A substantial number of monocytes/macrophages in inflamed appendiceal, synovial, and dermal tissues also reacted with antibody CSLEX1; however, only rare tissue macrophages in uninflamed nonlymphoid sites showed expression of SLex. These observations are consistent with the concept that SLex on circulating PMN and monocytes functions as a ligand for endothelial E-selectin in the development of inflammatory reactions. SLex-positive lymphocytes also were seen, notably, T lymphocytes in inflamed skin. An unexpected finding was that the CSLEX1 antibody also reacted with venular endothelium in certain lymphoid tissues and in inflamed appendix, but not with endothelium in normal appendix. Whether the SLex antigen identified on endothelium represents de novo expression or passive adsorption remains to be determined. 相似文献