Phospho-Smad2 (Ser 250) Rabbit mAb

Catalog No.: F3450

Application: Reactivity: Human, Mouse, Rat

Usage Information

Dilution
WB1:1000 IP1:30 IF1:50

Application
WB, IP, IF

Source
Rabbit

Reactivity
Human, Mouse, Rat

Storage Buffer
PBS, pH 7.2+50% Glycerol+0.05% BSA+0.01% NaN3

Storage (from the date of receipt)
-20°C (avoid freeze-thaw cycles), 2 years

Predicted MW
52 kDa

Positive Control	HeLa cell (treated with 0.2µM PMA for 0.5h); NIH/3T3 cell (treated with 0.2µM PMA for 0.5h); C6 cell (treated with 0.2µM PMA for 0.5h)
Negative Control

Datasheet & SDS

Biological Description

Specificity
Phospho-Smad2 (Ser 250) Rabbit mAb recognizes endogenous levels of total Smad2 protein only when phosphorylated at Ser 250.

Clone
J3P10

Synonym(s)
MADH2, MADR2, SMAD2, Mothers against decapentaplegic homolog 2, MAD homolog 2, Mothers against DPP homolog 2, JV18-1, Mad-related protein 2, SMAD family member 2, hMAD-2, SMAD 2, Smad2, Hsmad2

Background
Phosphorylation of Smad2 at serine 250 (pSmad2 S250), located in the linker region between the MH1 and MH2 domains, is a key regulatory modification mediated by kinases like nemo-like kinase (NLK). This phosphorylation acts as a negative regulator of canonical TGF-β signaling by inhibiting the critical C-terminal phosphorylation of Smad2, thus suppressing its transcriptional activity. Additionally, pSmad2 S250 promotes Smad2 ubiquitination and subsequent proteasomal degradation, providing a mechanism to fine-tune or terminate TGF-β responses. This modification is essential for controlling Smad2 stability and function under physiological conditions. Furthermore, pSmad2 S250 influences Smad2 nuclear translocation and affects downstream gene expression. Elevated levels of pSmad2 S250 are observed in proliferating cells, where it plays a role in cell cycle regulation and the modulation of TGF-β-dependent transcriptional programs.

Background

Phosphorylation of Smad2 at serine 250 (pSmad2 S250), located in the linker region between the MH1 and MH2 domains, is a key regulatory modification mediated by kinases like nemo-like kinase (NLK). This phosphorylation acts as a negative regulator of canonical TGF-β signaling by inhibiting the critical C-terminal phosphorylation of Smad2, thus suppressing its transcriptional activity. Additionally, pSmad2 S250 promotes Smad2 ubiquitination and subsequent proteasomal degradation, providing a mechanism to fine-tune or terminate TGF-β responses. This modification is essential for controlling Smad2 stability and function under physiological conditions. Furthermore, pSmad2 S250 influences Smad2 nuclear translocation and affects downstream gene expression. Elevated levels of pSmad2 S250 are observed in proliferating cells, where it plays a role in cell cycle regulation and the modulation of TGF-β-dependent transcriptional programs.

References
[1] Liang J, et al. J Biol Chem. 2021 Jan-Jun;296:100512. [2] Nitschkowski D, et al. Br J Cancer. 2025 May 3.

Reference Table for Selecting PVDF Membrane Pore Size Specifications

Protein Size (kDa)	PVDF Transfer Membrane Pore Size (μm)
< 10	0.22
10-20	0.22
20-30	0.45
30-45	0.45
45-70	0.45
80-100	0.45
100-140	0.45
> 140	0.45

Tech Support

Answers to questions you may have can be found in the inhibitor handling instructions. Topics include how to prepare stock solutions, how to store inhibitors, and issues that need special attention for cell-based assays and animal experiments.

Handling Instructions

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Protein Size (kDa)	Separating Gel Percentage
4-40	20%
12-45	15%
10-70	12.5%
15-100	10%
25-100	8%
60-210	5%