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Objective:To investigate the interference factors causing false-positive result of novel coronavirus IgM antibody detected by gold immunochromatography assay (GICA) and enzyme-linked immunosorbent assay (ELISA).Methods:A total of 71 serum from different pathogen infections and related chronic diseases patients were collected from the Affiliated Hospital of North Sichuan Medical College from January 22, 2020 to February 15, 2020. GICA and ELISA were used to detect 2019-nCoV IgM in 71 serum, including 5 influenza A virus (Flu A) IgM positive serum, 5 influenza B virus (Flu B) IgM positive serum, 5 Mycoplasma pneumonia (MP) IgM positive serum, 5 Legionella pneumophila (LP) IgM positive serum, 29 rheumatoid factor (RF) IgM positive serum, 5 hypertension patients serum, 5 diabetes mellitus patients serum, 6 human immunodeficiency virus (HIV) infection patients serum and 6 COVID-19 patients serum. The interference factors causing false positive results of the two methods were analyzed, and urea dissociation test was employed to dissociate the 2019-nCoV IgM positive serum using the best dissociation concentration. Statistical analyses were performed by SPSS, version 19.0.Results:2019-nCoV IgM was positive in 18 cases of middle-high level RF-IgM positive serum and 6 cases of 2019-nCoV-infected serum detected by two methods, and the other 47 serum were negative. When the dissociation concentration of urea was 6 mol/L, 2019-nCoV IgM was negative in 17 cases of middle-high level RF-IgM positive serum and positive in 6 cases of 2019-nCoV-infected serum detected by GICA. When the urea dissociation concentration was 4 mol/L, dissociation time was 10 min and the avidity index<0.46 was set as negative, 2019-nCoV IgM was negative in 15 cases of middle-high level RF-IgM positive serum and positive in 6 cases of 2019-nCoV-infected serum detected by ELISA.Conclusion:The middle-high level of RF-IgM could cause false positive results of 2019-nCoV IgM detected by GICA and ELISA, and the urea dissociation test would be helpful for reducing the probability of false-positive results of 2019-nCoV IgM test.
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Objective@#To investigate the interference factors causing false-positive result of novel coronavirus IgM antibody (SARS-CoV-2 IgM) detected by gold immunochromatography assay (GICA) and enzyme-linked immunosorbent assay (ELISA).@*Methods@#A total of 71 serum from different pathogen infections and related chronic diseases patients were collected from the Affiliated Hospital of North Sichuan Medical College from January 25, 2020 to February 15, 2020. GICA and ELISA were used to detect 2019-nCoV IgM in 71 serum, including 5 influenza A virus (Flu A) IgM positive serum, 5 influenza B virus (Flu B) IgM positive serum, 5 Mycoplasma pneumonia (MP) IgM positive serum, 5 Legionella pneumophila (LP) IgM positive serum, 29 rheumatoid factor (RF) IgM positive serum, 5 hypertension patients serum, 5 diabetes mellitus patients serum, 6 human immunodeficiency virus (HIV) infection patients serum and 6 Corona Virus Disease 2019 (COVID-19) patients serum. The interference factors causing false positive results of the two methods were analyzed, and urea dissociation test was employed to dissociate the 2019-nCoV IgM positive serum using the best dissociation concentration. Statistical analyses were performed by SPSS, version 19.0.@*Result@#s 2019-nCoV IgM was positive in 18 cases of middle-high level RF-IgM positive serum and 6 cases of 2019-nCoV-infected serum detected by two methods, and the other 47 serum were negative. When the dissociation concentration of urea was 6 mol/L, 2019-nCoV IgM was negative in 17 cases of middle-high level RF-IgM positive serum and positive in 6 cases of 2019-nCoV-infected serum detected by GICA. When the urea dissociation concentration was 4 mol/L, dissociation time was 10 min and the avidity index<0.46 was set as negative, 2019-nCoV IgM was negative in 15 cases of middle-high level RF-IgM positive serum and positive in 6 cases of 2019-nCoV-infected serum detected by ELISA.@*Conclusion@#The middle-high level of RF-IgM could cause false positive results of SARS-CoV-2 IgM detected by GICA and ELISA, and the urea dissociation test would be helpful for reducing the probability of false-positive results of SARS-CoV-2 IgM test.
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Objective To study the effect of TLR4 activation with LPS on BMP9-induced osteogenic differentiation of immortalized mouse embryonic fibroblasts ( iMEFs).Methods The activation of TLR4/NF-κB signaling path-way was detected by ICC.iMEFs were treated with LPS,BAY11-7082,Adnovirus GFP and BMP9.The early osteo-genic differentiation capability of iMEFs was detected by ALP staining and quantitative assay .The later osteogenic differentiation capability was detected by alizarin red S staining .The expression of later osteogenic differentiation marker gene OCN and OPN were detected by PCR and Western blot .The change of p-Smad1/5/8 was detected by Western blot.The expression of Runx2 and Dlx5 were detected by PCR and Western blot .Results LPS can effec-tively stimulate TLR4/NF-κB signaling pathway .TLR4 activation inhibited BMP 9-induced osteogenic differentiation . BMP9-induced osteogenic differentiation related gene and Smad 1/5/8 signaling activation were inhibited by TLR4 activation .The inhibition effect was partly reversed by BAY 11-7082 ( P<0.05 ) .Conclusions TLR4 activation with LPS can inhibit BMP9-induced osteogenic differentiation of iMEFs cells via NF-κB signaling pathway .
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AIM:To investigate the effect of bone morphogenetic proteins 9 (BMP9) on the migration and in-vasion abilities of human lung squamous-cell carcinoma NCI-H520 cells and its mechanism.METHODS:The expression of BMP9 at mRNA and protein levels in the NCI-H520 cells and human bronchial epithelial ( HBE) cells was detected by RT-PCR and Western blot.The NCI-H520 cells were transfected with the recombinant adenovirus AdBMP9 and the expres-sion of BMP9 at mRNA and protein levels was validated by RT-PCR and Western blot.The migration and invasion abilities of the NCI-H520 cells were determined by wound-healing and Transwell assays.The mRNA and protein levels of the migra-tion-related factor matrix metalloproteinase 2 (MMP2) were detected by RT-PCR and Western blot.The level of phospho-rylated Smad1/5 (p-Smad1/5) was detected by Western blot.Meanwhile, NCI-H520 cells were treated with BMP specific antagonist AdNoggin and AdBMP9.The level of p-Smad1/5 and the cell migration ability were measured by Western blot, wound-healing and Transwell assays.RESULTS:The expression of BMP9 at mRNA and protein levels was lower in NCI-H520 cells than that in HBE cells.After AdBMP9 was stably transfected into the NCI-H520 cells, the expression of BMP9 at mRNA and protein levels was significantly up-regulated, cell migration and invasion abilities were significantly de-creased, and the mRNA and protein levels of MMP2 were decreased.Meanwhile, the level of p-Smad1/5 was increased. Noggin reversed BMP9-caused the increase in p-Smad1/5 and the decrease in cell migration ability.CONCLUSION:O-ver-expression of BMP9 inhibits the migration and invasion abilities of lung squamous-cell carcinoma NCI-H520 cells.The activation of BMP-Smad signaling pathway may be involved in this inhibitory process.