The clinical presentation of COVID-19 and the specific antibody responses associated with SARS-Co... more The clinical presentation of COVID-19 and the specific antibody responses associated with SARS-CoV-2 variants have not been investigated during the emergence of Omicron variants in Bangladesh. The Delta and Omicron variants were identified by post-PCR melting curve analysis of the spike (S) protein receptor binding domain amplicons. Anti-S-protein immunoglobulin-G anti-nucleocapsid (N)-protein immunoglobulin-G and immunoglobulin-A levels were measured by ELISA. The Delta variant was found in 40 out of 40 (100%) SARS-CoV-2 RT-PCR positive COVID-19 patients between 13 September and 23 October 2021 and Omicron variants in 90 out of 90 (100%) RT-PCR positive COVID-19 patients between 9 January and 10 February 2022. The Delta variant associated with hospitalization (74%, 80%, and 40%) and oxygen support (60%, 57%, and 40%) in the no vaccine, dose-1, and dose-2 vaccinated cases, respectively, whereas the Omicron COVID-19 required neither hospitalization nor oxygen support (0%, p < 0.00...
Cathepsin G, a major proteolytic component of the neutrophil, acted on single-chain urokinase pla... more Cathepsin G, a major proteolytic component of the neutrophil, acted on single-chain urokinase plasminogen activator (sc-uPA) to generate a two chain form linked by disulphide bridges that was indistinguishable from plasmin generated tc-uPA by SDS-PAGE. Sequence analysis revealed that the cleavage site was located between 159 Ile and 160 Ile, as has previously been reported for elastase cleavage (Schmitt et al. 1989). A component (approximately 20%) was found to have been cleaved between 158 Lys and 159 Ile, the plasmin cleavage site and some plasminogen activating activity was found in the incubation mixture. A recombinant form of sc-uPA (rsc-uPA 158 Glu) was cleaved by cathepsin G but not plasmin. Cathepsin G action has also been studied in the presence of heparin, as a model for possible modes of interaction in the presence of the ECM. Cleavage to two chain form was mediated in a dose dependent form and there was an accumulation of a modified single chain form which migrated just ...
Objective: Autoantibodies to the thyrotropin hormone receptor (TSH-R) directly mediate Graves’ di... more Objective: Autoantibodies to the thyrotropin hormone receptor (TSH-R) directly mediate Graves’ disease (GD) and orbital manifestations of Graves’ orbitopathy (GO). To explore their heterogeneous function, TSH-R autoantibody binding in immunoassays (TRAb) versus their functional activity; stimulators of cAMP in bioassay (TSIs or TSAbs) or inhibitors of the thyrotropic hormone (TSH) binding to TSH-R, detected by decreased cAMP in bioassay (TBAbs), is assessed.Methods: TRAb measurements by automated electrochemical luminescence immunoassays (ECLIA) and stimulating and blocking activity on Chinese hamster ovary cell (CHO) Mc4 bioassays (Thyretain ®, Quidel). The activity of patient immunoglobulins on ECLIA; M22 Mab competition (cobas elecsys, Roche) or TSH-R bridge assay (Immulite,Siemens) versus Mc4 TSI bioassay were correlated with GO clinical activity by meta-analysis of complete Pubmed-published data sets (n=5).Results: The automated ECLIA show less sensitivity than cell-based bioas...
Background: The ophthalmopathy associated with Graves’ hyperthyroidism is most likely a T-cell me... more Background: The ophthalmopathy associated with Graves’ hyperthyroidism is most likely a T-cell mediated disorder, although most investigators believe that autoantobodies directed against the TSH Receptor (TSH-R) expressed on the surface of the orbital fibroblasts and pre-adipocytes are the main driver of the orbital reactions. Cases of ophthalmopathy without TSH-R binding antibodies and not closely associated with Thyroid Stimulating Immunoglobulin’s (TSI), challenge the notion that ophthalmopathy is perpetrated by TSI circulating in peripheral blood and TSI producing plasma cells resident in the orbital tissues. One way to address a possible role of TSH-R antibodies in the development of ophthalmopathy is to study patients with so-called “Euthyroid Graves’ eye Disease (EGD)”, who have a similar eye disorder but with normal thyroid function and no features of thyroid autoimmunity during long term follow up. Methods: The cell-based reporter bioassay Thyretain TM was used to assess TS...
Background: The assessment of antibody responses to severe acute respiratory syndrome coronavirus... more Background: The assessment of antibody responses to severe acute respiratory syndrome coronavirus-2 is potentially confounded by exposures to flaviviruses. The aims of the present research were to determine whether anti-dengue antibodies affect the viral load and the detection of anti-coronavirus nucleocapsid (N)-protein antibodies in coronavirus infectious disease 2019 (COVID-19) in Bangladesh. Methods: Viral RNA was evaluated in swab specimens from 115 COVID-19 patients by real-time reverse transcription polymerase chain reaction (rT-PCR). The anti-N-protein antibodies, anti-dengue virus E-protein antibodies and the dengue non-structural protein-1 were determined in serum from 115 COVID-19 patients, 30 acute dengue fever pre-COVID-19 pandemic and nine normal controls by ELISA. Results: The concentrations of viral RNA in the nasopharyngeal; Ct median (95% CI); 22 (21.9–23.3) was significantly higher than viral RNA concentrations in oropharyngeal swabs; and 29 (27–30.5) p < 0.000...
The aim of this study was to investigate the potential of the new TSH-receptor antibody (TRAb) as... more The aim of this study was to investigate the potential of the new TSH-receptor antibody (TRAb) assays to predict remission or relapse of hyperthyroidism in patients with Graves’ disease (GD) and Graves’ orbitopathy (GO). TRAbs were measured retrospectively in sera from a cohort of GD patients with GO (n=117; remission n=38 and relapse n=79–Essen GO biobank) with automated binding immunoassays: TRAb Elecsys (Cobas Roche) and TRAb bridge assay (IMMULITE, Siemens), and the TSAb (thyroid stimulating Ab) cell-based bioassay (Thyretain, Quidel Corp.). To identify relapse risk/remission of hyperthyroidism patients were followed up at least 10 months after the end of antithyroid drug therapy (ATD) therapy. ROC plot analysis was performed to calculate cut-off levels of TRAb and TSAb for prediction of relapse and remission of hyperthyroidism. Cut-off serum levels are provided for timepoints around 3, 6, 10, and 15 months after the beginning of ATD. Repeated measurements of TRAb increase the r...
We have previously shown that polyclonal rabbit ATG from Fresenius (ATG-F) can induce non-overt D... more We have previously shown that polyclonal rabbit ATG from Fresenius (ATG-F) can induce non-overt DIC in patients undergoing hematopoietic stem cell transplantation (HSCT) [Weber et al. 2003]. The compensated state of the coagulopathy suggested that the severe drop in platelet counts (>60%) observed in patients with normal pretreatment levels was not solely due to thrombin generation and platelet consumption. We thus further investigated the mechanisms of ATG-F-induced thrombocytopenia, hypothesizing that ATG-F had direct platelet-stimulating activity. Using an indirect flow cytometric binding assay we found that, compared to control IgG, ATG-F dose-dependently (1–100 μg/ml) bound to the surface of resting and TRAP-activated platelets, showing an up to 50fold increase in MFI at 50–100 μg/ml. In a washed platelet system, in which baseline positivity for CD62P was generally…
Proceedings of the National Academy of Sciences, 2004
Complex recombinant antibody fragments for modulation of immune function such as tumor cell destr... more Complex recombinant antibody fragments for modulation of immune function such as tumor cell destruction have emerged at a rapid pace and diverse anticancer strategies are being developed to benefit patients. Despite improvements in molecule design and expression systems, the quantity and stability, e.g., of single-chain antibodies produced in cell culture, is often insufficient for treatment of human disease, and the costs of scale-up, labor, and fermentation facilities are prohibitive. The ability to yield mg/ml levels of recombinant antibodies and the scale-up flexibility make transgenic production in plants and livestock an attractive alternative to mammalian cell culture as a source of large quantities of biotherapeutics. Here, we report on the efficient production of a bispecific single-chain antibody in the serum of transgenic rabbits and a herd of nine cloned, transgenic cattle. The bispecific protein, designated r28M, is directed to a melanoma-associated proteoglycan and the...
The ability of Leishmania mexicana amazonensis to inhibit antigen specific T-cell proliferation a... more The ability of Leishmania mexicana amazonensis to inhibit antigen specific T-cell proliferation against a non-parasite polypeptide antigen, poly(LTyr, LGlu)-poly(DLAla)--poly(LLys), was examined. Infection of mouse peritoneal macrophages by promastigotes blocked the proliferation of the T-cell line, TPB1. This effect was correlated with the level of parasite infection, and the timing of macrophage infection and antigen addition. Peritoneal macrophages from both BALB/b and C57BL/6 mice showed reduced ability to serve as antigen presenting cells.
An isolate of Beet black scorch virus (BBSV) was obtained from Iranian sugar beet roots. Its geno... more An isolate of Beet black scorch virus (BBSV) was obtained from Iranian sugar beet roots. Its genome organization closely resembles that of the previously described Chinese and North American isolates, but the nucleotide sequences of the three isolates differ considerably. Most of the nucleotide exchanges, however, are silent, and the Iranian and the Chinese isolates were serologically indistinguishable. Beets infected by the Iranian BBSV did not show black scorch symptoms, but severe root beardedness. This might have been caused by BBSV or the simultaneously present beet necrotic yellow vein virus, or both together.
The aim of the study was to investigate the use of serial measurements of TSH-receptor autoantibo... more The aim of the study was to investigate the use of serial measurements of TSH-receptor autoantibodies (TRAb) with the newest available assay technology to predict the course of Graves’ Orbitopathy (GO) during the first 24 months from disease onset. Serial serum samples from patients with GO (103 mild/135 severe) were collected between 2007 and 2017 and retrospectively analyzed. The course of GO were classified into mild/severe 12 months after manifestation (severe: NOSPECS≥5; mild<5). TRAb were measured with automated binding immunoassays (IU/l): TRAb Elecsys (Cobas, Roche), TRAb bridge assay (IMMULITE, Siemens), and a cell-based bioassay (percent of specimen to reference ratio - SRR%) (Thyretain, Quidel). Variable cut off levels of measured TRAb were calculated at specificity of 90% from receiver operator curve (ROC) analysis for several timepoints during the course of GO. To select one: 5–8 months after first GO symptoms, which is the timepoint for usual referals for treatment ...
The clinical presentation of COVID-19 and the specific antibody responses associated with SARS-Co... more The clinical presentation of COVID-19 and the specific antibody responses associated with SARS-CoV-2 variants have not been investigated during the emergence of Omicron variants in Bangladesh. The Delta and Omicron variants were identified by post-PCR melting curve analysis of the spike (S) protein receptor binding domain amplicons. Anti-S-protein immunoglobulin-G anti-nucleocapsid (N)-protein immunoglobulin-G and immunoglobulin-A levels were measured by ELISA. The Delta variant was found in 40 out of 40 (100%) SARS-CoV-2 RT-PCR positive COVID-19 patients between 13 September and 23 October 2021 and Omicron variants in 90 out of 90 (100%) RT-PCR positive COVID-19 patients between 9 January and 10 February 2022. The Delta variant associated with hospitalization (74%, 80%, and 40%) and oxygen support (60%, 57%, and 40%) in the no vaccine, dose-1, and dose-2 vaccinated cases, respectively, whereas the Omicron COVID-19 required neither hospitalization nor oxygen support (0%, p < 0.00...
Cathepsin G, a major proteolytic component of the neutrophil, acted on single-chain urokinase pla... more Cathepsin G, a major proteolytic component of the neutrophil, acted on single-chain urokinase plasminogen activator (sc-uPA) to generate a two chain form linked by disulphide bridges that was indistinguishable from plasmin generated tc-uPA by SDS-PAGE. Sequence analysis revealed that the cleavage site was located between 159 Ile and 160 Ile, as has previously been reported for elastase cleavage (Schmitt et al. 1989). A component (approximately 20%) was found to have been cleaved between 158 Lys and 159 Ile, the plasmin cleavage site and some plasminogen activating activity was found in the incubation mixture. A recombinant form of sc-uPA (rsc-uPA 158 Glu) was cleaved by cathepsin G but not plasmin. Cathepsin G action has also been studied in the presence of heparin, as a model for possible modes of interaction in the presence of the ECM. Cleavage to two chain form was mediated in a dose dependent form and there was an accumulation of a modified single chain form which migrated just ...
Objective: Autoantibodies to the thyrotropin hormone receptor (TSH-R) directly mediate Graves’ di... more Objective: Autoantibodies to the thyrotropin hormone receptor (TSH-R) directly mediate Graves’ disease (GD) and orbital manifestations of Graves’ orbitopathy (GO). To explore their heterogeneous function, TSH-R autoantibody binding in immunoassays (TRAb) versus their functional activity; stimulators of cAMP in bioassay (TSIs or TSAbs) or inhibitors of the thyrotropic hormone (TSH) binding to TSH-R, detected by decreased cAMP in bioassay (TBAbs), is assessed.Methods: TRAb measurements by automated electrochemical luminescence immunoassays (ECLIA) and stimulating and blocking activity on Chinese hamster ovary cell (CHO) Mc4 bioassays (Thyretain ®, Quidel). The activity of patient immunoglobulins on ECLIA; M22 Mab competition (cobas elecsys, Roche) or TSH-R bridge assay (Immulite,Siemens) versus Mc4 TSI bioassay were correlated with GO clinical activity by meta-analysis of complete Pubmed-published data sets (n=5).Results: The automated ECLIA show less sensitivity than cell-based bioas...
Background: The ophthalmopathy associated with Graves’ hyperthyroidism is most likely a T-cell me... more Background: The ophthalmopathy associated with Graves’ hyperthyroidism is most likely a T-cell mediated disorder, although most investigators believe that autoantobodies directed against the TSH Receptor (TSH-R) expressed on the surface of the orbital fibroblasts and pre-adipocytes are the main driver of the orbital reactions. Cases of ophthalmopathy without TSH-R binding antibodies and not closely associated with Thyroid Stimulating Immunoglobulin’s (TSI), challenge the notion that ophthalmopathy is perpetrated by TSI circulating in peripheral blood and TSI producing plasma cells resident in the orbital tissues. One way to address a possible role of TSH-R antibodies in the development of ophthalmopathy is to study patients with so-called “Euthyroid Graves’ eye Disease (EGD)”, who have a similar eye disorder but with normal thyroid function and no features of thyroid autoimmunity during long term follow up. Methods: The cell-based reporter bioassay Thyretain TM was used to assess TS...
Background: The assessment of antibody responses to severe acute respiratory syndrome coronavirus... more Background: The assessment of antibody responses to severe acute respiratory syndrome coronavirus-2 is potentially confounded by exposures to flaviviruses. The aims of the present research were to determine whether anti-dengue antibodies affect the viral load and the detection of anti-coronavirus nucleocapsid (N)-protein antibodies in coronavirus infectious disease 2019 (COVID-19) in Bangladesh. Methods: Viral RNA was evaluated in swab specimens from 115 COVID-19 patients by real-time reverse transcription polymerase chain reaction (rT-PCR). The anti-N-protein antibodies, anti-dengue virus E-protein antibodies and the dengue non-structural protein-1 were determined in serum from 115 COVID-19 patients, 30 acute dengue fever pre-COVID-19 pandemic and nine normal controls by ELISA. Results: The concentrations of viral RNA in the nasopharyngeal; Ct median (95% CI); 22 (21.9–23.3) was significantly higher than viral RNA concentrations in oropharyngeal swabs; and 29 (27–30.5) p < 0.000...
The aim of this study was to investigate the potential of the new TSH-receptor antibody (TRAb) as... more The aim of this study was to investigate the potential of the new TSH-receptor antibody (TRAb) assays to predict remission or relapse of hyperthyroidism in patients with Graves’ disease (GD) and Graves’ orbitopathy (GO). TRAbs were measured retrospectively in sera from a cohort of GD patients with GO (n=117; remission n=38 and relapse n=79–Essen GO biobank) with automated binding immunoassays: TRAb Elecsys (Cobas Roche) and TRAb bridge assay (IMMULITE, Siemens), and the TSAb (thyroid stimulating Ab) cell-based bioassay (Thyretain, Quidel Corp.). To identify relapse risk/remission of hyperthyroidism patients were followed up at least 10 months after the end of antithyroid drug therapy (ATD) therapy. ROC plot analysis was performed to calculate cut-off levels of TRAb and TSAb for prediction of relapse and remission of hyperthyroidism. Cut-off serum levels are provided for timepoints around 3, 6, 10, and 15 months after the beginning of ATD. Repeated measurements of TRAb increase the r...
We have previously shown that polyclonal rabbit ATG from Fresenius (ATG-F) can induce non-overt D... more We have previously shown that polyclonal rabbit ATG from Fresenius (ATG-F) can induce non-overt DIC in patients undergoing hematopoietic stem cell transplantation (HSCT) [Weber et al. 2003]. The compensated state of the coagulopathy suggested that the severe drop in platelet counts (>60%) observed in patients with normal pretreatment levels was not solely due to thrombin generation and platelet consumption. We thus further investigated the mechanisms of ATG-F-induced thrombocytopenia, hypothesizing that ATG-F had direct platelet-stimulating activity. Using an indirect flow cytometric binding assay we found that, compared to control IgG, ATG-F dose-dependently (1–100 μg/ml) bound to the surface of resting and TRAP-activated platelets, showing an up to 50fold increase in MFI at 50–100 μg/ml. In a washed platelet system, in which baseline positivity for CD62P was generally…
Proceedings of the National Academy of Sciences, 2004
Complex recombinant antibody fragments for modulation of immune function such as tumor cell destr... more Complex recombinant antibody fragments for modulation of immune function such as tumor cell destruction have emerged at a rapid pace and diverse anticancer strategies are being developed to benefit patients. Despite improvements in molecule design and expression systems, the quantity and stability, e.g., of single-chain antibodies produced in cell culture, is often insufficient for treatment of human disease, and the costs of scale-up, labor, and fermentation facilities are prohibitive. The ability to yield mg/ml levels of recombinant antibodies and the scale-up flexibility make transgenic production in plants and livestock an attractive alternative to mammalian cell culture as a source of large quantities of biotherapeutics. Here, we report on the efficient production of a bispecific single-chain antibody in the serum of transgenic rabbits and a herd of nine cloned, transgenic cattle. The bispecific protein, designated r28M, is directed to a melanoma-associated proteoglycan and the...
The ability of Leishmania mexicana amazonensis to inhibit antigen specific T-cell proliferation a... more The ability of Leishmania mexicana amazonensis to inhibit antigen specific T-cell proliferation against a non-parasite polypeptide antigen, poly(LTyr, LGlu)-poly(DLAla)--poly(LLys), was examined. Infection of mouse peritoneal macrophages by promastigotes blocked the proliferation of the T-cell line, TPB1. This effect was correlated with the level of parasite infection, and the timing of macrophage infection and antigen addition. Peritoneal macrophages from both BALB/b and C57BL/6 mice showed reduced ability to serve as antigen presenting cells.
An isolate of Beet black scorch virus (BBSV) was obtained from Iranian sugar beet roots. Its geno... more An isolate of Beet black scorch virus (BBSV) was obtained from Iranian sugar beet roots. Its genome organization closely resembles that of the previously described Chinese and North American isolates, but the nucleotide sequences of the three isolates differ considerably. Most of the nucleotide exchanges, however, are silent, and the Iranian and the Chinese isolates were serologically indistinguishable. Beets infected by the Iranian BBSV did not show black scorch symptoms, but severe root beardedness. This might have been caused by BBSV or the simultaneously present beet necrotic yellow vein virus, or both together.
The aim of the study was to investigate the use of serial measurements of TSH-receptor autoantibo... more The aim of the study was to investigate the use of serial measurements of TSH-receptor autoantibodies (TRAb) with the newest available assay technology to predict the course of Graves’ Orbitopathy (GO) during the first 24 months from disease onset. Serial serum samples from patients with GO (103 mild/135 severe) were collected between 2007 and 2017 and retrospectively analyzed. The course of GO were classified into mild/severe 12 months after manifestation (severe: NOSPECS≥5; mild<5). TRAb were measured with automated binding immunoassays (IU/l): TRAb Elecsys (Cobas, Roche), TRAb bridge assay (IMMULITE, Siemens), and a cell-based bioassay (percent of specimen to reference ratio - SRR%) (Thyretain, Quidel). Variable cut off levels of measured TRAb were calculated at specificity of 90% from receiver operator curve (ROC) analysis for several timepoints during the course of GO. To select one: 5–8 months after first GO symptoms, which is the timepoint for usual referals for treatment ...
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