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Biomedical subjects

A Mulder

Publications and source records attributed to A Mulder.

66 records · Page 4Linked to original sources
PubMed1989

Rapid detection of human cytomegalovirus DNA in peripheral blood leukocytes of viremic transplant recipients by the polymerase chain reaction.

Peripheral blood leukocyte samples (n = 458) of 24 bone marrow transplant and 52 kidney transplant patients were examined weekly for the presence of human cytomegalovirus (HCMV) using an improved culture technique (DEAFF; detection of early antigen fluorescent foci). In total 5 (21%) bone marrow transplant and 11 (21%) kidney transplant patients developed a viremia. Patients' samples were investigated for the presence of HCMV DNA using an in vitro DNA amplification technique, the polymerase chain reaction (PCR). From the statistically evaluable viremic patients (n = 13), 110 blood samples were analyzed. In 5 of these patients, the DEAFF and PCR led to identical results. In 8 patients however the PCR was more sensitive, i.e. HCMV DNA was detected for a longer period of time. Applying statistical analysis using the McNemar test, this result was significant (P less than 0.05). The PCR applied on leukocyte samples did not detect HCMV DNA in viruric patients without viremia. Moreover, the current PCR never led to positive results with peripheral blood leukocyte samples of healthy seropositive or seronegative controls. Since the PCR can be performed in 6 hr, this technique will contribute to rapid detection of HCMV DNA in peripheral blood leukocytes and therefore to optimal clinical management of HCMV-infected transplant recipients.

Antigens, Viral↗
PubMed1989

Detection of cytomegalovirus antigens and DNA in tissues fixed in formaldehyde.

Immunohistochemical techniques with monoclonal antibodies against cytomegalovirus (CMV) immediate early (IEA) and early antigens (EA), and in situ hybridisation, were used to detect CMV infection in routinely obtained, formaldehyde fixed and paraffin wax embedded tissues taken from bone marrow transplant patients, who had died form interstitial pneumonia. To improve the rates of detection of CMV-IEA and EA the wax embedded material was pretreated with 0.4% pepsin/HCl at 37 degrees C for 30 minutes. This pretreatment was also advantageous for in situ hybridisation. In the patients with histological evidence of CMV infection or positive viral culture from the lung tissue, or both, viral proteins and nucleic acids were detected in lung, as well as in other organs. Immunohistochemical techniques proved superior in heavily infected but necrotic tissues. In control patients (patients who had died from interstitial pneumonia without any evidence of CMV, or with no interstitial pneumonia at all) in situ hybridisation showed no positive signal, while immunohistochemical techniques showed only a few positive cells in lung tissue of one of nine patients. In addition to CMV-DNA analysis, formaldehyde-fixed, paraffin wax embedded tissue is amenable to immunohistochemical analysis with CMV monoclonal antibodies.

Antigens, Viral↗
PubMed1989

An IgM capture enzyme linked immunosorbent assay to detect IgM antibodies to treponemes in patients with syphilis.

A new IgM capture enzyme linked immunosorbent assay (ELISA) was compared with the 19S(IgM) fluorescent treponemal antibody absorption (19S(IgM)FTA-ABS) test for detecting IgM antibodies to treponemes. Serum samples from 180 people, 109 with various stages of untreated syphilis, 45 with treated syphilis, and 26 non-infected, were investigated. In all diagnostic groups of syphilis the reactivity of the IgM capture ELISA was similar to that of the 19S(IgM)FTA-ABS test except in untreated neurosyphilis, for which the IgM capture ELISA was significantly less sensitive. The IgM capture ELISA was very sensitive in congenital (100%, 5/5) and primary (82%, 18/22) syphilis, but less sensitive in secondary (60%, 12/20), latent (53%, 16/30), neurosyphilis (34%, 11/32), and treated (11%, 5/45) syphilis. False positive IgM capture ELISA results were not found in five people who gave false positive Venereal Disease Research Laboratory (VDRL) reactions or in 21 neonates born to mothers adequately treated for syphilis before or during pregnancy. This indicated that the IgM capture ELISA was very specific. The course of antitreponemal IgM reactivity after treatment of early infectious syphilis was followed up in six patients. The quantity of IgM antibody declined in nearly all patients after treatment, but still remained detectable in five patients up to six months after treatment. In contrast, non-treponemal antibodies measured by the VDRL test disappeared in four out of six patients within five months from starting treatment. In conclusion, the IgM capture ELISA may be useful for easy and sensitive detection of IgM antibodies to treponemes in patients with congenital and primary syphilis. A positive test result in these cases indicates that patients should receive treatment if they have not been treated recently. The test is not, however, recommended to replace the VDRL test to monitor patients treated for syphilis.

Antibodies, Bacterial↗
PubMed1984

Myeloid-associated antigen 3-alpha-fucosyl-N-acetyllactosamine (FAL): location on various granulocyte membrane glycoproteins and masking upon monocytic differentiation.

Seven different granulocyte-reactive murine monoclonal antibodies (mAb) were studied. The antigens recognized by these mAb were immunoprecipitated from lysates of 125I-labeled granulocytes of healthy donors. The isolated antigens were analyzed by electrophoresis on sodium dodecyl sulfate-polyacrylamide gel and autoradiography. All 7 antibodies precipitated the same 6 membrane polypeptides from membrane-iodinated granulocyte lysates: 105 and 150-kDa as most pronounced, together with 260-, 230-, 67- and 52-kDa polypeptides. One of the antibodies studied, B4.3, is directed against 3-alpha-fucosyl-N-acetyllactosamine as shown by absorption with the synthesized carbohydrate molecule. Competition experiments with 125I-labeled B4.3 demonstrated complete inhibition of binding by B4.3 and 3 of the other antibodies (VM D5, UJ308, MI/N1) and partial inhibition by the 3 other antibodies (FMC 10, FMC 12, FMC 13), indicating binding to the same antigenic structure. None of the 7 mAb reacted with monocytes in the immunofluorescence technique, but after neuraminidase treatment of these cells, positive reactions were obtained with all mAb. Immunoprecipitation with lysates of both native and neuraminidase-treated monocytes showed no polypeptide bands. Monocytic differentiation of the cell line HL60 by 12-O-tetradecanoylphorbol-13-acetate (TPA) and of cell line U 937 by dimethylsulfoxide and TPA was accompanied by a decrease in reactivity with these antibodies, which could be recovered by neuraminidase treatment. This indicates that 3-alpha-fucosyl-N-acetyllactosamine is masked for the detection of the antibody upon monocytic differentiation by sialylation.

Antibodies, Monoclonal↗
PubMed1984

Megakaryoblastic differentiation of proerythroblastic K562 cell-line cells.

The human proerythroblastic leukemia cell-line K562 was induced to differentiate into megakaryocytic cells by 12-O-tetradecanoylphorbol-13-acetate (TPA). Megakaryocytic differentiation was detected when lineage-specific monoclonal antibodies were used to monitor the effect of TPA on K562 cells. A monoclonal anti-platelet antibody (C17) directed against an epitope present on GP IIIa appeared to react with K562 cells after induction. This was observed together with the disappearance of glycophorin A, the erythrocyte-specific lineage antigen. The induced megakaryocytic cells were also detected by ultrastructural platelet peroxidase (PPO). Immunoprecipitation, after ectolabeling of the cells with the C17 antibody and SDS-polyacrylamide gel electrophoresis, proved that TPA-induced K562 expressed both GP IIIa and GP IIb. However, the monoclonal antibody C15 directed against another epitope of platelet GP IIIa reacted only partially, or not at all, indicating that GP IIIa expressed on TPA-induced K562 differs structurally from that on normal platelets. K562 clones, expressing glycophorin A in all cells, were obtained by limiting dilution and culture. When these clones were treated with TPA, again megakaryocytic cells were obtained. These findings are discussed in relation to normal megakaryocytopoiesis.

Butyrates↗
PubMed1984

Immunochemical characterization of platelet-specific alloantigens.

Immunoprecipitation was performed with platelet-specific alloantisera (anti-Zwa, -Zwb, -Baka and antiserum Luc) and 125I-labelled platelets of a panel of donors typed for these platelet-specific alloantigens. This was done by sensitization of intact, radiolabelled platelets with the antisera, solubilization of the sensitized platelets with Nonidet P-40 and recovery of the immune complexes from the lysate with heat-killed, formalin-fixed Staphylococcus aureus, strain Cowan I. In the case of platelet antibodies of the IgM class, bacteria also preincubated with rabbit-IgG anti-human IgM before treatment of the lysate were applied. Radiolabelled antigens were then eluted from the bacteria by boiling in an SDS-mercaptoethanol-containing buffer. Subsequently, the eluted antigens were analyzed by SDS-polyacrylamide gel electrophoresis. It revealed that both the Zwa and the Baka antigens are located on the glycoprotein-IIb/IIIa complex. The location of the Zwb antigen could not be established in this way. From the intensity of the precipitated glycoprotein bands, it appeared that Zwa is probably a marker of glycoprotein IIIa, Baka or glycoprotein IIb. However, immunoprecipitates of anti-Zwa and anti-Baka sera both carried, to some extent, complexes of glycoproteins IIb and IIIa. Iodinated platelets of a Glanzmann thrombasthenia patient failed to show any material precipitated by these antisera or the antiserum Luc, known to contain antibodies against both glycoproteins.

Blood Platelets↗
PubMed1982

Seasonality of pre-ovulatory non-disjunction and the aetiology of Down syndrome. A European collaborative study.

Six series of patients with Down syndrome (DS) from different European countries, altogether 287 cases, were divided into four categories according to parental origin of the additional chromosome 21 and meiotic division in which the nondisjunction had occurred. The monthly birth or conception frequencies per category were analysed by graph and compared with the total birth curve by Watson's adaptation of the Kolmogorov-Smirnov statistic for cyclic trends. Unexpectedly, the non-disjunctions during maternal meiosis I (63%), by far the largest category, occurred more frequently during the seasonal "restoration" and "inhibition" phase of the "ovulatory seasons" and less frequently when the ovulation rate is stabilized. The graph of the maternal meiosis II patients (17%) also seemed to conform to this phenomenon, though less obviously. In contrast to this, the paternal DS graph (20%) was very divergent, although a seasonal cluster of non-disjunctions may also occur here. From these findings a seasonal disturbance of preovulatory ripening of the ovum emerges as a possible cause of the first (and second) meiotic non-disjunction. Seasonal periodicity of the prolactin concentration in women and "transient hyperprolactinaemia", shown to be allied to delayed ovulation, may be related to these seasonal DS conception clusters.

Down Syndrome↗
PubMed1981

Characterization, by immunoprecipitation, of myeloid- and monocyte-specific antigens present on the human promyelocytic cell line (HL-60) in three stages of differentiation.

The human promyelocytic leukemia cell line HL-60 is reactive with an antiserum raised against normal human granulocytes (AGS). Immunoprecipitation with AGS on [35S]methionine-labeled HL-60 cell lysates with subsequent analysis by NaDodSO4/polyacrylamide slab gel electrophoresis shows a major antigenic doublet with molecular weights of 88,000 and 86,000, together with some minor antigens of lower molecular weight. Upon stimulation with dimethyl sulfoxide or 12-O-tetradecanoylphorbol 13-acetate, which induces HL-60 to differentiate to mature granulocytes or monocytes/macrophages, respectively, this antigenic doublet disappears. 12-O-Tetradecanoylphorbol 13-acetate induces the synthesis of an antigen, molecular weight 83,000, reactive with an antimonocyte serum. Neutrophil-specific alloantigens were not detected on HL-60 or its differentiated derivatives.

Animals↗
PubMedDate not supplied

Reactivity of twenty-two cytotoxic human monoclonal HLA antibodies towards soluble HLA class I in an enzyme-linked immunosorbent assay (PRA-STAT).

An ELISA, PRA-STAT was recently introduced for the detection of HLA class I specific antibodies of IgG isotype in patients' sera. We studied the antigenicity of the soluble HLA (sHLA) preparations that are used in this ELISA as the detection matrix, with the aid of a panel of complement binding human HLA monoclonal antibodies (HuMAbs). A total of 22 HuMAbs, including both IgG and IgM were used. CDC and PRA-STAT ELISA were in complete agreement on 9 of the mAbs tested, with 16 HLA-A and 16 HLA-B locus antigens or their splits identified identically on CDC and PRA-STAT. In 7 of the remaining 13 HuMAbs, there was a difference of one antigen in the specificity pattern of the two techniques three times a specificity call not made by CDC, and four times a call not made by PRA-STAT. For the remaining 6 HuMAbs the differences involve 2 antigens (4 HuMAbs), and 3 or 4 antigens (1 HuMAb each). This study shows the validity of PRA-STAT for detection of HLA-class I antibodies, irrespective of isotype, in serum. The immunological integrity of the sHLA preparations used in PRA-STAT is also confirmed, albeit with some slight discrepancies in antibody specificity seen between PRA-STAT and CDC.

Antibodies, Monoclonal↗