Biomedical subjects

S Kuijper

Publications and source records attributed to S Kuijper.

29 records · Page 2Linked to original sources

PubMed1993

Evaluation of different tests for the serodiagnosis of tuberculosis and the use of likelihood ratios in serology.

A serologic test for the diagnosis of tuberculosis was evaluated in 91 newly diagnosed tuberculosis (TB) patients, of whom 15 were HIV positive, in 17 TB patients during treatment, and in 220 control subjects (including individuals from endemic areas and patients with sarcoidosis or Crohn's disease). Purified proteins of M. tuberculosis with molecular weights of 10,000, 16,000, 24,000, 30,000, 38,000, and 70,000 were tested by ELISA. In addition, monoclonal antibody TB72 was tested by competition ELISA. The cutoff values were set at the mean plus three standard deviations of the values obtained in 100 healthy Dutch army recruits. Only the ELISA with the 10,000, 16,000, and 24,000 antigens and the TB72 assay discriminated between patients with TB who were not HIV positive and control subjects. Specificity varied from 95 to 98% and sensitivity from 29 to 51% with the different antigens. Combination of the test results of the ELISA with the 16,000 antigen and the TB72 assay had a sensitivity of 65% (95% confidence interval, 53 to 75%) and a specificity of 96% (95% confidence interval, 92 to 98%). The assay was useful for the diagnosis of both pulmonary and extrapulmonary TB. Optimal use of the serologic assay could be obtained when likelihood ratios for each test value are calculated instead of using the test dichotomized (positive or negative). High posttest probabilities indicate the presence of TB; low posttest probabilities do not exclude the disease and should lead to additional investigations.

AIDS-Related Opportunistic Infections↗

PubMed1992

Antibodies against secreted and non-secreted antigens in mice after infection with live Mycobacterium tuberculosis.

Mice from four different inbred strains were infected with live Mycobacterium tuberculosis and the immune response to M. tuberculosis was followed for 24 weeks, using Western blotting. Nearly all mice, irrespective of H-2 type, reacted with the 38-kDa protein band. Antibodies against this secreted 38-kDa protein were the first to appear, 4 weeks after infection. Thereafter the secreted 19-kDa protein and non-secreted antigens, such as the 65-kDa and 33-kDa proteins, were recognized. The immune response against the non-secreted antigens was influenced by the mouse strain. However, the 33-kDa protein band was recognized by all mouse strains after a second injection with live M. tuberculosis. The specificity of the antibodies was analysed in Western blot using sonicates of M. tuberculosis, M. kansasii, M. avium, M. terrae, M. gordonae and Escherichia coli. Antibodies against the 38-kDa and 33-kDa protein bands seemed to be specific for M. tuberculosis, while antibodies against the 19-kDa protein band showed limited cross-reactivity. Antibodies against the 65-kDa protein were strongly cross-reactive. These results suggest that the 38-kDa protein is secreted in vivo and, therefore, may be available to the humoral immune system at an early stage of infection. The non-secreted 33-kDa protein is only recognized by all mouse strains after prolonged contact with M. tuberculosis.

Animals↗

PubMed1992

Results of the third immunology of leprosy/immunology of tuberculosis antimycobacterial monoclonal antibody workshop.

An international workshop was sponsored by the World Health organization to screen new antimycobacterial monoclonal antibodies and to identify antibodies which could be recommended as standard reagents giving consistent results under differing assay conditions. Fifty-eight antibodies were submitted to the workshop by eight independent laboratories. Nineteen of the antibodies recognized antigens distinct from those identified in earlier workshops, defining at least 10 new protein antigens. Monoclonal antibodies characterized in the workshop provide a set of convenient reagents for further characterization of mycobacterial antigens.

Animals↗

PubMed1992

Detection of Mycobacterium tuberculosis in clinical samples by using polymerase chain reaction and a nonradioactive detection system.

A test based on the polymerase chain reaction (PCR) was developed for the detection of the Mycobacterium tuberculosis complex in clinical samples. In this test, a 245-bp sequence of the insertion element IS986 was amplified and detected by agarose gel electrophoresis in the presence of ethidium bromide and by Southern blot and dot blot hybridization by using a 188-bp digoxigenin-labeled probe. We tested clinical specimens from 227 patients suspected of having tuberculosis. These included 102 cerebrospinal fluid, 48 sputum, 18 pleural fluid, 5 bronchoalveolar lavage, 18 blood, 7 pus, 8 bone marrow, and 6 urine samples and 15 tissue biopsy specimens. We also tested sputum samples from 75 patients with diseases other than tuberculosis. Sputum samples were first decontaminated, and all samples were treated with proteinase K-detergent solution to extract the DNA. Part of each sample was spiked with M. tuberculosis to provide a semiquantitative assay and to control for the loss of mycobacteria or interference with the PCR which may cause false-negative results. One femtogram of M. tuberculosis DNA could be detected. PCR was positive for all 32 culture-positive (for M. tuberculosis) and Ziehl-Neelsen staining (ZN)-positive samples, 10 of 12 culture-positive and ZN-negative samples, and all 4 culture-negative and ZN-positive samples. PCR detected M. tuberculosis complex bacteria in 35 of 178 culture- and ZN-negative samples. Clinical data supported the diagnosis of tuberculosis in the majority of the 35 patients from whom those samples were obtained.

Base Sequence↗

PubMed1991

[Development of a serological test for tuberculosis].

Antigens secreted by M. tuberculosis in the culture medium and antigens obtained from sonicated M. tuberculosis were characterized, at the laboratory of Tropical Hygiene of the Royal Tropical Institute and the division of Pulmonary Medicine at the Academic Medical Centre, in order to explore which antigens could be valuable in the development of a serological test for tuberculosis. Using murine monoclonal antibodies in immunoblot, a pattern of protein bands distinct from that of the corresponding M. tuberculosis sonicate was found in the culture medium. The major protein bands of the culture medium, of 24 and 12 kD, and the major protein band of 16 kD in sonicate were purified. These antigens were tested in ELISA with sera from 20 patients with tuberculosis, diagnosed by a positive culture of M. tuberculosis, and from 21 control subjects. The ELISA results obtained with these 3 antigens were combined and the mean value obtained in the control group plus 2 times the standard deviation was chosen as the cut-off level. Sixteen of the 20 patients with tuberculosis had antibodies against 1 of the purified antigens, while none of the control subjects had. By combining the results, obtained with these 3 antigens, 17 of the 20 patients with tuberculosis were positive in this serological test and none of the control subjects. The 24, 12 and 16 kD antigens may be valuable for the development of a serological test for tuberculosis.

Antigens, Bacterial↗

PubMed1990

Antigens in culture supernatant of Mycobacterium tuberculosis: epitopes defined by monoclonal and human antibodies.

Antigens of Mycobacterium tuberculosis found in the supernatant of heat-treated cultures were characterized in order to explore whether antigens from this source could be used for the development of a serological test. Culture supernatants and sonicates of 12, 25 and 39 d cultures were analysed by SDS-PAGE. In culture supernatant, major protein bands of 65, 24, and 12 kDa were visible after Coomassie brilliant blue staining. Using murine monoclonal antibodies in Western blots, a pattern of protein bands distinct from that of the corresponding M. tuberculosis sonicates was found in all the culture supernatants. Gel permeation chromatography, in the presence of SDS, was used to separate the major protein bands in the culture supernatant. In ELISA, sera from 20 of 26 patients with tuberculosis reacted with fractions containing mainly 24 kDa or 12 kDa proteins, whereas none of the control sera reacted. In Western blots, each patient serum had its own characteristic banding pattern with culture supernatant, but all the sera from tuberculosis patients and control subjects reacted with protein bands of 65, 61, 58, 30 and 24 kDa. The 12 kDa protein was recognized only by sera from patients with tuberculosis in both Western blots and ELISA. This suggests that different kinds of epitopes on proteins of M. tuberculosis are detected by human antibodies in Western blots and ELISA. We assume that epitopes recognized in Western blots by patients with tuberculosis and control subjects are ubiquitous and are also present on normal commensal bacteria. Epitopes recognized by only some patients with tuberculosis in Western blots may be linear and M. tuberculosis specific. Epitopes recognized by tuberculosis patients but by none of the control subjects in ELISA may be conformation related and M. tuberculosis specific. The major protein bands found in supernatants of heat-treated cultures, 24 and 12 kDa, possess epitopes that may be M. tuberculosis specific and are potentially valuable for the development of a serological test.

Antibodies, Bacterial↗

PubMed1990

Anti-Mycobacterium leprae monoclonal antibodies cross-react with human skin: an alternative explanation for the immune responses in leprosy.

A panel of 17 mouse monoclonal antibodies (MoAb) raised against Mycobacterium leprae (M. leprae) antigens was used to detect antigenic determinants in normal human skin. An indirect immunoperoxidase technique was used. Eight of the MoAb detected epidermal antigens similar to patterns well known for human sera. Five of these MoAb detected determinants in the dermis, too. These observations may indicate a certain degree of similarity between the antigenic determinants occurring in M. leprae and in the human host. We propose that such a similarity on the one hand may facilitate the survival of M. leprae in the human host when the antigens are not recognized as "non-self," a situation which seems to occur in lepromatous leprosy, when the patients' tissues are loaded with bacteria virtually without any immune response. On the other hand, M. leprae antigens which mimic host antigens may induce an auto-immune reaction against the host's own antigens, which could explain the immune reaction in tuberculoid leprosy and during a "reversal reaction" when M. leprae is not observed in the host tissues, but extensive granuloma formation occurs.

Animals↗

PubMed1990

Enzyme immunoassay for identification of heat-killed mycobacteria belonging to the Mycobacterium tuberculosis and Mycobacterium avium complexes and derived from early cultures.

A simple enzyme-linked immunosorbent assay was developed for the identification of cultured mycobacteria belonging to the Mycobacterium tuberculosis complex, the Mycobacterium avium complex, and Mycobacterium kansasii. Six monoclonal antibodies were used: two (F23-49 and F24-2) were specific for the M. tuberculosis complex, two (F85-2 and F85-10) were specific for the identification of the M. avium complex, one (F126-22) was specific for M. kansasii, and one (F141-3) was broadly reactive and distinguished mycobacteria from other bacteria. In the enzyme-linked immunosorbent assay only 10(6) to 10(7) bacteria derived from early cultures (2 to 3 weeks) were needed for each monoclonal antibody. For the M. tuberculosis complex the sensitivity and specificity were both 100%. For the M. avium complex the specificity was 100% and the sensitivity was 70%. The three M. kansasii strains tested could all be identified as M. kansasii.

Antibodies, Monoclonal↗

PubMed1989

Production and characterization of monoclonal antibodies against specific serotypes of Mycobacterium avium and the Mycobacterium avium-Mycobacterium intracellulare-Mycobacterium scrofulaceum complex.

Serotype-specific and Mycobacterium avium-Mycobacterium intracellulare-Mycobacterium scrofulaceum complex (MAIS complex)-specific monoclonal antibodies (MAbs) were prepared. A series of MAbs were obtained, five specific for serotype 2, three specific for serotype 4, eight against a strain of serotype 19, two specific for the MAIS complex, and two against a common glycolipid shared by all the mycobacteria tested so far. The serotype-specific and the MAIS complex-specific MAbs reacted in immunofluorescence with intact mycobacteria and in enzyme-linked immunosorbent assay and immuno-thin-layer chromatography with lipid extracts of mycobacteria. The two MAbs against a common mycobacterial glycolipid reacted only in lipid enzyme-linked immunosorbent assay and immuno-thin-layer chromatography. All MAbs were directed against glycopeptidolipids (GPLs), except for four MAbs against proteins of serotype 19. The serotype- and MAIS complex-specific epitopes on GPLs are exposed on the mycobacterial cell wall, in contrast with the common mycobacterial glycolipid, which is probably located inside the cell wall. The serotype-specific MAbs reacted with native as well as deacetylated GPLs, in contrast with the MAIS complex-specific MAbs, which reacted only with native GPLs. The MAbs will be useful for the identification of MAIS complex and M. avium serotypes 2 and 4 and a strain of serotype 19, GPL analyses with immuno-thin-layer chromatography, and the localization of GPL epitopes in mycobacteria.

Animals↗

PubMed1984

Production and characterization of monoclonal antibodies to Mycobacterium tuberculosis, M. bovis (BCG) and M. leprae.

Thirty-two monoclonal antibodies (MoAb) to Mycobacterium tuberculosis H37Rv, M. bovis BCG and M. leprae were produced. The spleen cells of BALB/c mice immunized with sonicated or intact bacilli were fused with Sp2/0-Ag-14 myeloma cells. Many more antibody producing hybridomas were found when M. tuberculosis, rather than M. leprae, was used as the immunogen. The MoAb were characterized by an enzyme immunoassay and immunofluorescence on 16 mycobacterial species. The sodium dodecylsulphate polyacrylamide gel electrophoresis immunoperoxidase assay was used to determine the molecular weight of the antigens detected by the MoAb. Antigens of high, low and intermediate molecular weight were found. Some of the antigens were proteinaceous, others of a glycolipid nature. The immunofluorescence assay proved to be essential for the selection of MoAb since some MoAb reacted only in this assay and not in the enzyme immunoassay. The most specific clones were found in the fusions with spleen cells of mice immunized with intact rather than sonicated bacteria. One MoAb (F29-29) reacted only with M. tuberculosis H37Rv; one (F41-3) only with M. leprae and another (F29-45) reacted with M. tuberculosis and M. gastrii. Several MoAb only reacted with three mycobacterial species: M. tuberculosis, M. kansasii and M. gastrii. Others showed unique patterns of reactivity by enzyme immuno- and immunofluorescence assay. The potential use of the MoAb for the identification of mycobacteria and mycobacterial antigens is discussed.

Animals↗

PubMedDate not supplied

Rapid identification of cultured Mycobacterium tuberculosis with a panel of monoclonal antibodies in western blot and immunofluorescence.

This paper describes the identification of cultured mycobacteria with a panel of monoclonal antibodies directed against species-specific epitopes in a Western blot (WB) test and in an immunofluorescence test (IFT). In WB, we identified mycobacteria of the Mycobacterium tuberculosis complex (M. tuberculosis, M. africanum, M. bovis, M. bovis BCG, and M. microti) with 10(8) bacteria. In the IFT, we identified mycobacteria of the M. tuberculosis complex, the M. avium complex (M. avium, M. intracellulare and M. scrofulaceum) and M. kansasii with 10(7) bacteria. Using a panel of 105 mycobacterial patient isolates, we compared identification by WB and IFT with conventional, culture and biochemical identification. Identification of the M. tuberculosis complex in WB had a specificity and sensitivity of 96.3 and 98.3%, respectively. Identification in IFT of the M. tuberculosis complex had a specificity and sensitivity of 94.6% and 89.7%, respectively. Since the identification of mycobacteria with mAb requires only a small number of bacteria, these tests will reduce by several weeks the time necessary for microbiological identification.

Antibodies, Monoclonal↗