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Phospho-BTK (Tyr223) Antibody [H3N9]

Cat.No.: F0977

Application: Reactivity: Human

Usage Information

Dilution
WB1:1000 IP1:20

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 Observed MW
76 kDa 76 kDa
^*Why do the predicted and actual molecular weights differ? The following reasons may explain differences between the predicted and actual protein molecular weight.

Datasheet & SDS

Biological Description

Specificity
Phospho-BTK (Tyr223) Antibody [H3N9] detects endogenous levels of total BTK protein only when it is phosphorylated at Tyr223.

Clone
H3N9

Synonym(s)
AGMX1; ATK; BPK; BTK; Tyrosine-protein kinase BTK; Agammaglobulinemia tyrosine kinase; B-cell progenitor kinase; Bruton tyrosine kinase

Background
Phospho-BTK (Tyr223) denotes the activated state of Bruton's tyrosine kinase, a TEC family non-receptor tyrosine kinase essential for B-cell receptor signaling and maturation. This protein possesses a pleckstrin homology (PH) domain for PIP3-driven membrane localization, followed by TEC homology, SH3, SH2, and kinase domains. Activation is initiated by Src family kinases such as Lyn phosphorylating BTK at Tyr551 in the kinase domain, which then enables autophosphorylation at Tyr223 within the SH3 domain. This sequential phosphorylation stabilizes BTK’s open, active conformation, boosting kinase activity toward downstream targets like PLCγ2, which in turn drives IP3 and DAG production, calcium flux, and activation of NF-κB and MAPK cascades. These signals are crucial for B-cell proliferation, differentiation, survival, and effective antigen presentation, while PKCβ-mediated phosphorylation at Ser180 provides negative feedback to set signaling thresholds. Persistent phosphorylation at Tyr223 is implicated in B-cell malignancies such as chronic lymphocytic leukemia and lymphoma, making BTK a major target for therapeutics like ibrutinib. Loss-of-function mutations in BTK, meanwhile, result in X-linked agammaglobulinemia.

Background

Phospho-BTK (Tyr223) denotes the activated state of Bruton's tyrosine kinase, a TEC family non-receptor tyrosine kinase essential for B-cell receptor signaling and maturation. This protein possesses a pleckstrin homology (PH) domain for PIP3-driven membrane localization, followed by TEC homology, SH3, SH2, and kinase domains. Activation is initiated by Src family kinases such as Lyn phosphorylating BTK at Tyr551 in the kinase domain, which then enables autophosphorylation at Tyr223 within the SH3 domain. This sequential phosphorylation stabilizes BTK’s open, active conformation, boosting kinase activity toward downstream targets like PLCγ2, which in turn drives IP3 and DAG production, calcium flux, and activation of NF-κB and MAPK cascades. These signals are crucial for B-cell proliferation, differentiation, survival, and effective antigen presentation, while PKCβ-mediated phosphorylation at Ser180 provides negative feedback to set signaling thresholds. Persistent phosphorylation at Tyr223 is implicated in B-cell malignancies such as chronic lymphocytic leukemia and lymphoma, making BTK a major target for therapeutics like ibrutinib. Loss-of-function mutations in BTK, meanwhile, result in X-linked agammaglobulinemia.

References
https://pubmed.ncbi.nlm.nih.gov/9326643/ https://pubmed.ncbi.nlm.nih.gov/9391111/

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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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%