Western blot analysis of extracts from SK-N-MC cells, untreated (-) or treated with Forskolin #3828 (30 μM) and IBMX (0.5 mM) for 30 min (+), using Phospho-CREB (Ser133) (87G3) Rabbit mAb #9198 (upper) and CREB (48H2) Rabbit mAb #9197 (lower).
Boil for 3 minutes prior to use. Load 10 µl of phosphorylated and nonphosphorylated CREB Control Cell Extracts per lane.
Supplied in SDS Sample Buffer: 62.5 mM Tris-HCL (pH 6.8 at 25°C), 2% w/v SDS, 10% glycerol, 50 mM DTT, 0.01% w/v bromophenol blue or phenol red. Store at –20°C.
CREB Control Cell Extracts (SK-N-MC untreated): Total cell extracts from SK-N-MC cells serve as a negative control. Supplied in SDS sample buffer.
CREB Control Cell Extracts (SK-N-MC +IBMX/Forskolin): Total cell extracts from SK-N-MC cells treated with 30 μM Forskolin #3828 and 0.5 mM IBMX for 30 minutes serve as a positive control. Supplied in SDS sample buffer.
CREB is a bZIP transcription factor that activates target genes through cAMP response elements. CREB is able to mediate signals from numerous physiological stimuli, resulting in regulation of a broad array of cellular responses. While CREB is expressed in numerous tissues, it plays a large regulatory role in the nervous system. CREB is believed to play a key role in promoting neuronal survival, precursor proliferation, neurite outgrowth, and neuronal differentiation in certain neuronal populations (1-3). Additionally, CREB signaling is involved in learning and memory in several organisms (4-6). CREB is able to selectively activate numerous downstream genes through interactions with different dimerization partners. CREB is activated by phosphorylation at Ser133 by various signaling pathways including Erk, Ca2+, and stress signaling. Some of the kinases involved in phosphorylating CREB at Ser133 are p90RSK, MSK, CaMKIV, and MAPKAPK-2 (7-9).
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