Phospho-IRAK4 (Thr345/Ser346) Antibody [G2K20]

Application: Reactivity: Human

Usage Information

Dilution
WB1:1000 IP1:50

Application
WB, IP

Reactivity
Human

Source
Rabbit Monoclonal Antibody

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
55 kDa

Positive Control	KARPAS-299 cells (hIL-1β, 50 ng/ml, 15 min)
Negative Control	KARPAS-299 cells (serum-starved)

Datasheet & SDS

Biological Description

Specificity
Phospho-IRAK4 (Thr345/Ser346) Antibody [G2K20] detects endogenous levels of total IRAK4 protein only when it is phosphorylated at Thr345/Ser346.

Clone
G2K20

Synonym(s)
Interleukin-1 receptor-associated kinase 4; IRAK-4; Renal carcinoma antigen NY-REN-64; IRAK4

Background
Phospho-IRAK4 (Thr345/Ser346) is the activated form of the serine/threonine kinase IRAK4, a key mediator of innate immune signaling through IL-1R and Toll-like receptors (TLRs). IRAK4 contains an N-terminal death domain for MyD88 binding and a C-terminal kinase domain with essential activation loop residues (Thr342, Thr345, Ser346, Thr352). Upon receptor stimulation, MyD88 recruits IRAK4, triggering dimerization and intermolecular autophosphorylation, primarily at Thr345/Ser346, which is necessary for full kinase activation. Phosphorylation of any two among Thr342, Thr345, or Ser346 is sufficient. This phosphorylation, which occurs 15–30 minutes after IL-1 or TLR agonist stimulation and is blocked by IRAK4 inhibitors, enables IRAK4 to phosphorylate downstream targets like Pellino1 and IRAK1, leading to Myddosome formation and activation of TAK1, IKKs (NF-κB pathway), and MAPKs (JNK, p38, ERK), resulting in cytokine production (e.g., IL-6, TNF-α). The functional impact of phospho-IRAK4 is cell-type specific: in primary human monocytes, kinase inhibition reduces cytokine output, while in dermal fibroblasts, IRAK4’s scaffolding function is sufficient for signaling, though both functions are lost in IRAK4-deficient cells. Dysregulated IRAK4 activity is implicated in autoimmune and inflammatory diseases, with kinase-dead mutants offering protection in models such as arthritis; while IRAK4 has some basal activity, it becomes strongly hyperphosphorylated upon immune activation.

Background

Phospho-IRAK4 (Thr345/Ser346) is the activated form of the serine/threonine kinase IRAK4, a key mediator of innate immune signaling through IL-1R and Toll-like receptors (TLRs). IRAK4 contains an N-terminal death domain for MyD88 binding and a C-terminal kinase domain with essential activation loop residues (Thr342, Thr345, Ser346, Thr352). Upon receptor stimulation, MyD88 recruits IRAK4, triggering dimerization and intermolecular autophosphorylation, primarily at Thr345/Ser346, which is necessary for full kinase activation. Phosphorylation of any two among Thr342, Thr345, or Ser346 is sufficient. This phosphorylation, which occurs 15–30 minutes after IL-1 or TLR agonist stimulation and is blocked by IRAK4 inhibitors, enables IRAK4 to phosphorylate downstream targets like Pellino1 and IRAK1, leading to Myddosome formation and activation of TAK1, IKKs (NF-κB pathway), and MAPKs (JNK, p38, ERK), resulting in cytokine production (e.g., IL-6, TNF-α). The functional impact of phospho-IRAK4 is cell-type specific: in primary human monocytes, kinase inhibition reduces cytokine output, while in dermal fibroblasts, IRAK4’s scaffolding function is sufficient for signaling, though both functions are lost in IRAK4-deficient cells. Dysregulated IRAK4 activity is implicated in autoimmune and inflammatory diseases, with kinase-dead mutants offering protection in models such as arthritis; while IRAK4 has some basal activity, it becomes strongly hyperphosphorylated upon immune activation.

References
https://pubmed.ncbi.nlm.nih.gov/28512203/ https://pubmed.ncbi.nlm.nih.gov/24567333/

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

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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%