The mitogen-activated protein kinase (MAPK) signaling pathway is a typical example of a protein kinase cascade. MAPKs generally form multi-tiered pathways that receive input from several levels above. Typically, a MAPK is activated through phosphorylation by an activated MAP2K, which is activated through phosphorylation by an activated MAP3K. Unlike MAP3Ks, the activation mechanisms of MAPKs and MAP2Ks are relatively simple. Invitrogen MAPK signaling pathway antibodies are designed to dependably detect the key MAPK targets. Each antibody is validated for use in various applications.
Key MAPK targets include
Invitrogen MAPK signaling pathway antibodies are designed to dependably detect the key MAPK targets.
Immunofluorescent analysis of ERK3 (green) showing positive staining in the cytoplasm and nucleus of HT-29 cells (right) compared with a negative control in the absence of primary antibody (left). Formalin-fixed cells were permeabilized with 0.1% Triton X-100 in TBS for 5-10 minutes, blocked with 3% BSA-PBS for 30 minutes at room temperature and probed with an ERK3 monoclonal antibody (Cat. No. MA1-101) in 3% BSA-PBS at a dilution of 1:200 and incubated overnight at 4 °C in a humidified chamber. Cells were washed with PBST and incubated with a DyLight 488-conjugated goat-anti-mouse IgG (H+L) secondary antibody in PBS at room temperature in the dark. F-actin (red) was stained with a fluorescent red phalloidin and nuclei (blue) were stained with DAPI for 5-10 minutes in the dark. Images were taken at a magnification of 60x.
Immunohistochemistry analysis of ERK1/2 (pTpY185/187) showing staining in the cytoplasm and nucleus of paraffin-embedded human colon carcinoma tissue (right) compared to a negative control without primary antibody (left). To expose target proteins, antigen retrieval was performed using 10mM sodium citrate (pH 6.0), microwaved for 8-15 min. Following antigen retrieval, tissues were blocked in 3% H2O2-methanol for 15 min at room temperature, washed with ddH2O and PBS, and then probed with a ERK1/2 (pTpY185/187) polyclonal antibody (Cat. No. 44-680G) diluted in 3% BSA-PBS at a dilution of 1:20 overnight at 4°C in a humidified chamber. Tissues were washed extensively in PBST and detection was performed using an HRP-conjugated secondary antibody followed by colorimetric detection using a DAB kit. Tissues were counterstained with hematoxylin and dehydrated with ethanol and xylene to prep for mounting.
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