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Abcam’s SMAD1 (pS463/S465) in vitro SimpleStep ELISA™ (Enzyme-Linked Immunosorbent Assay) kit is designed for the semi-quantitative measurement of SMAD1 (pS463/S465) protein in human and mouse cells.
The SimpleStep ELISA™ employs a labeled capture and detector antibody which immunocaptures the sample analyte in solution. This entire complex (capture antibody/protein/detector antibody) is in turn immobilized in the well by immunoaffinity via the anti-tag antibody. Samples or standards are added to the wells, followed by the antibody mix. After incubation, the wells are washed to remove unbound material; the TMB substrate is then added. The reaction is stopped by addition of Stop Solution which stops the color development and completes any color change from blue to yellow. Signal is generated proportionally to the amount of bound analyte and the intensity is measured at 450 nm. Optionally, instead of the endpoint reading, development of TMB can be recorded kinetically at 600 nm.
|Components||1 x 96 tests|
|10X Wash Buffer PT||1 x 15ml|
|50X Cell Extraction Enhancer Solution||1 x 1ml|
|5X Cell Extraction Buffer PTR (ab193970)||1 x 12ml|
|Lyophilized SMAD1 (pS463/S465) Control Lysate||1 vial|
|Plate Seal||1 unit|
|SimpleStep Pre-Coated 96-Well Microplate (ab206978)||1 unit|
|SMAD1 (pS463/S465) Capture Antibody||1 x 3ml|
|SMAD1 (pS463/S465) Detector Antibody||1 x 3ml|
|Stop Solution||1 x 12ml|
|TMB Substrate||1 x 12ml|
Our Abpromise guarantee covers the use of ab186036 in the following tested applications.
The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.
|Sandwich ELISA||Use at an assay dependent concentration.|
Example of a typical SMAD1 (pS463/S465) cell lysate dilution series. Raw data values are shown in the table. Background-subtracted data values (mean +/- SD) are graphed.
Linearity of dilution in representative sample matrices. Cellular lysates were prepared at 3 concentrations in common media containing 1X Cell Extraction Buffer PTR. Data from duplicate measurements of SMAD1 (pS463/S465) are normalized and plotted.
Cell line analysis for Total SMAD1 from 300 µg/mL preparations of cell extracts. Data from triplicate measurements (mean +/- SD) are plotted and compared to 1X Cell Extraction Buffer PTR (zero).
Induction of SMAD1 (pS463/S465) phosphorylation in HeLa cells in response to BMP-4 treatment. HeLa cells were cultured in 96-well tissue culture plates, serum-starved and treated (30 min) with a dose-range of BMP-4 before cell lysis. Data from quadruplicate measurements of SMAD1 (pS463/S465) are plotted.
Inhibition of SMAD1 (pS463/S465) phosphorylation in HeLa cells in response to dorsomorphin treatment. HeLa cells were cultured in 96-well tissue culture plates and treated with a dose-range of dorsomorphin (30 min). Cells were then stimulated with BMP-4 (30 min) and lysed. Data from quadruplicate measurements of SMAD1 (pS463/S465) and SMAD1 (Total) are plotted.
ab186036 has not yet been referenced specifically in any publications.