Western blot analysis of purified MAPK phospho-proteins or extracts from NIH/3T3 cells treated with UV light and PDGF, using Phospho-p44/42 MAPK (Thr202/Tyr204) (197G2) Rabbit mAb (upper), Phospho-p38 MAPK (Thr180/Tyr182) (3D7) Rabbit mAb #9215 (middle), and Phospho-SAPK/JNK (Thr183/Tyr185) (98F2) Rabbit mAb #4671 (lower).
Western blot analysis of extracts from 293 cells and NIH/3T3 cells, untreated or UV-treated, using Phospho-SAPK/JNK (Thr183/Tyr185) (98F2) Rabbit mAb (upper) or SAPK/JNK (56G8) Rabbit mAb (lower).
|REACTIVITY||H M R Hm|
|MW (kDa)||46, 54|
Supplied in 10 mM sodium HEPES (pH 7.5), 150 mM NaCl, 100 µg/ml BSA, 50% glycerol and less than 0.02% sodium azide. Store at –20°C. Do not aliquot the antibody.
For western blots, incubate membrane with diluted primary antibody in 5% w/v BSA, 1X TBS, 0.1% Tween® 20 at 4°C with gentle shaking, overnight.
NOTE: Please refer to primary antibody datasheet or product webpage for recommended antibody dilution.
From sample preparation to detection, the reagents you need for your Western Blot are now in one convenient kit: #12957 Western Blotting Application Solutions Kit
NOTE: Prepare solutions with reverse osmosis deionized (RODI) or equivalent grade water.
Load 20 µl onto SDS-PAGE gel (10 cm x 10 cm).
NOTE: Volumes are for 10 cm x 10 cm (100 cm2) of membrane; for different sized membranes, adjust volumes accordingly.
* Avoid repeated exposure to skin.
posted June 2005
revised November 2013
Reprobing of an existing membrane is a convenient means to immunoblot for multiple proteins independently when only a limited amount of sample is available. It should be noted that for the best possible results a fresh blot is always recommended. Reprobing can be a valuable method but with each reprobing of a blot there is potential for increased background signal. Additionally, it is recommended that you verify the removal of the first antibody complex prior to reprobing so that signal attributed to binding of the new antibody is not leftover signal from the first immunoblotting experiment. This can be done by re-exposing the blot to ECL reagents and making sure there is no signal prior to adding the next primary antibody.
NOTE: Prepare solutions with reverse osmosis deionized (RODI) or equivalently purified water.
posted June 2005
revised October 2016
Protocol Id: 10
Phospho-SAPK/JNK (Thr183/Tyr185) (98F2) Rabbit mAb detects endogenous levels of p46 and p54 SAPK/JNK only when phosphorylated at Thr183 and Tyr185. This antibody does not recognize phosphorylated p44/42 MAPK or p38 MAP kinase.Species Reactivity:
Human, Mouse, Rat, Hamster
Monoclonal antibody is produced by immunizing animals with a synthetic phosphopeptide corresponding to residues surrounding Thr183/Tyr185 of human SAPK/JNK.
The stress-activated protein kinase/Jun-amino-terminal kinase SAPK/JNK is potently and preferentially activated by a variety of environmental stresses including UV and gamma radiation, ceramides, inflammatory cytokines, and in some instances, growth factors and GPCR agonists (1-6). As with the other MAPKs, the core signaling unit is composed of a MAPKKK, typically MEKK1-MEKK4, or by one of the mixed lineage kinases (MLKs), which phosphorylate and activate MKK4/7. Upon activation, MKKs phosphorylate and activate the SAPK/JNK kinase (2). Stress signals are delivered to this cascade by small GTPases of the Rho family (Rac, Rho, cdc42) (3). Both Rac1 and cdc42 mediate the stimulation of MEKKs and MLKs (3). Alternatively, MKK4/7 can be activated in a GTPase-independent mechanism via stimulation of a germinal center kinase (GCK) family member (4). There are three SAPK/JNK genes each of which undergoes alternative splicing, resulting in numerous isoforms (3). SAPK/JNK, when active as a dimer, can translocate to the nucleus and regulate transcription through its effects on c-Jun, ATF-2, and other transcription factors (3,5).
Explore pathways + proteins related to this product.
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U.S. Patent No. 7,429,487, foreign equivalents, and child patents deriving therefrom.