goat anti smad1 polyclonal antibody  (R&D Systems)

Journal: Cell death & disease

Article Title: F-box protein FBXO30 mediates retinoic acid receptor γ ubiquitination and regulates BMP signaling in neural tube defects.

doi: 10.1038/s41419-019-1783-y

Figure Lengend Snippet: Fig. 5 FBXO30 antagonizes the effect of RA on BMP signaling. a HEK293 cells were transfected overexpression of FBXO30. Thirty-six hours after transfection, BRE-luciferase activity was measured after treatments with BMP-2 (100 ng/ml) and/or RA (1 μM) for 12 h. Data are mean ± s.d. (n = 3). b HEK293 cells were transfected with the siRNA-FBXO30. Thirty-six hours after transfection, BRE-luciferase activity was measured after treatments with BMP-2 (100 ng/ml) and/or RA (1 μM) for 12 h. Data are mean ± s.d. (n = 3). c, d Overexpression of FBXO30 or siRNA-FBXO30 transfected in HEK293 cells as indicated. Thirty-six hours after transfection, cells were stimulated with BMP-2 and/or RA for 12 h before harvesting. Western blot was performed to examine the pSmad1/5 and total Smad1/5 levels in whole-cell lysates. Aliquots of total lysates were immunoblotted to indicate antibody. Numbers at the bottom were generated by quantification (Image J) of the pSmad1/5 signal normalized to the Smad1/5 signal. e siRNA- FBXO30 or siRNA-normal transfected in HEK293 cells as indicated. Thirty-six hours after transfection, cells were stimulated with BMP-2 and/or RA for 12 h before harvesting. Direct visualization or indirect immunofluorescence was performed. Scale bars, 22 μm

Article Snippet: Incubation with antibodies against FBXO30 (1:50 dilution; Santa-Cruz), p-smad1/5 (1:100; CST) was performed at 4 °C for 18 h. Quality assessment was performed on each batch of slides by including a negative control in which the primary antibody was replaced by 10% normal goat serum to preclude nonspecific signals.

Techniques: Transfection, Over Expression, Luciferase, Activity Assay, Western Blot, Generated