Customize ASK3 Antibody

Description

ASK3 Antibody Overview

ASK3 antibodies target the mitogen-activated protein kinase kinase kinase 15 (MAP3K15), a 147.4 kDa protein with 1,313 amino acids and three isoforms . These antibodies enable:

Detection of ASK3 in biological samples via Western blot (WB), ELISA, and immunohistochemistry (IHC) .
Investigation of ASK3's role in protein phosphorylation and stress-response pathways .

Commercial Availability
Nine ASK3 antibody products are available across suppliers like MyBioSource.com, with applications spanning WB, ELISA, flow cytometry (FCM), and IHC . A subset is highlighted below:

Product Name	Applications	Reactivity	Supplier
MAP3K15 Antibody (N-term)	WB, ELISA, FCM, IHC	Human, Mouse	MyBioSource.com

ASK3 in Osmotic Stress and Blood Pressure Regulation

ASK3 bidirectionally responds to osmotic stress (hypotonic and hypertonic conditions) and suppresses the WNK1-SPAK/OSR1 signaling pathway in renal tubules .

Mechanism: ASK3 inhibits WNK1 kinase activity, reducing phosphorylation of SPAK/OSR1 and downstream Na⁺-Cl⁻ cotransporter (NCC) activation .
Phenotypic Evidence:
- Ask3 knockout mice exhibit hypertension and hyperactivated SPAK/OSR1 in renal tubules .
- ASK3 localizes to the apical lumen of renal epithelial cells, suggesting a direct role in ion homeostasis .

Proteasomal Regulation of ASK3 Activity

Under hyperosmotic stress, the 19S proteasome subunit PSMC2 negatively regulates ASK3 activity.

Experimental Validation:
- PSMC2 depletion via siRNA increases ASK3 phosphorylation (Thr808), enhancing kinase activity .
- Immunofluorescence assays using phospho-specific antibodies confirm ASK3 activation dynamics .

Methodological Applications

ASK3 antibodies are pivotal in advanced experimental techniques:

Immunofluorescence-Based Activity Assays: Quantify ASK3 activation using phospho-specific antibodies (e.g., Thr808 phosphorylation) .
High-Content Screening: Genome-wide siRNA screens coupled with ASK3 antibody staining identify proteasomal regulators of ASK3 .

Implications for Hypertension Research

The interplay between ASK3 and the WNK1-SPAK/OSR1 pathway offers therapeutic targets for hypertension. ASK3 antibodies facilitate:

Tracking NCC activation in renal tissues.
Validating in vivo models of blood pressure dysregulation .

Product Specs

Buffer

Preservative: 0.03% Proclin 300
Constituents: 50% Glycerol, 0.01M PBS, pH 7.4

Form

Liquid

Lead Time

Made-to-order (14-16 weeks)

Synonyms

ASK3 antibody; SK12 antibody; At3g05840 antibody; F10A16.14Shaggy-related protein kinase gamma antibody; EC 2.7.11.1 antibody; ASK-gamma antibody; Shaggy-related protein kinase 12 antibody; AtSK12 antibody

Target Names

ASK3

Uniprot No.

P43289

Target Background

Function

ASK3 antibody may mediate extracellular signals to regulate transcription in differentiating cells. It is likely involved in two key processes: first, at the cortical polarity site, to restrict MAPK signaling and promote asymmetric cell division (ACD); and second, in the nucleus of stomatal lineage ground cells (SLGCs) or meristemoids, to limit cell division and promote differentiation into pavement or stomatal guard cells, respectively. ASK3 antibody is known to phosphorylate YDA and SPCH in vitro.

Gene References Into Functions

Research findings indicate that AtSK12 mediates brassinosteroid-regulated cell growth but not stomatal development, while BIN2 regulates both processes. PMID: 24292946

Database Links

KEGG: ath:AT3G05840

STRING: 3702.AT3G05840.1

UniGene: At.132

Protein Families

Protein kinase superfamily, CMGC Ser/Thr protein kinase family, GSK-3 subfamily

Subcellular Location

Cytoplasm. Cytoplasm, cell cortex.

Tissue Specificity

Roots, shoots and leaves.

Q&A

Here’s a structured FAQ for ASK3 antibody research, synthesized from peer-reviewed studies and technical guidelines:

Advanced Research Questions

What methods are recommended for investigating ASK3’s role in cellular stress pathways?

Functional Assays:
- Kinase Activity: Measure ATP consumption or Syntide 2 phosphorylation in vitro .
- Osmotic Stress Models: Expose renal tubular cells to hypertonic media and monitor ASK3-WNK1-SPAK/OSR1 pathway activation via phospho-specific antibodies .
- Apoptosis Assays: Use TNF-α or anti-Fas antibody treatments in ASK3 knockdown HeLa cells .

How can I resolve cross-reactivity issues in ASK3 immunofluorescence?

Sequence Alignment: Compare ASK3 epitopes with homologous proteins (e.g., ASK1/ASK2) to identify unique regions .
Peptide Blocking: Preincubate antibodies with excess immunizing peptide to confirm signal specificity .

What protocols optimize ASK3 detection in co-immunoprecipitation experiments?

Native Conditions: Use non-denaturing lysis buffers (e.g., 1% NP-40) to preserve protein interactions .
Protease Inhibition: Include phosphatase inhibitors to maintain phosphorylation states (critical for ASK3-WNK1 binding studies) .

Table 1: ASK3 Antibody Validation Checklist

Parameter	Recommendation	Reference
Epitope	Prefer N-terminal (e.g., MAP3K15 Antibody)
Application	WB: Denatured antigen; IHC: Native antigen
Specificity Control	siRNA knockdown + blocking peptide
Tissue Localization	Prioritize kidney tubule lysates

Table 2: ASK3 Functional Roles in Disease Models

Model System	Key Finding	Mechanism
HeLa Cell Knockdown	Reduced cytotoxicity under TNF-α/Fas stress	ASK3-MAPK pathway inhibition
Ask3 KO Mice	Hypertension + SPAK/OSR1 hyperactivation	Dysregulated WNK1 signaling
Cirrhosis Liver	↑ ASK3 expression vs. healthy tissue	Stress-responsive kinase

Technical Notes

AI-Driven Epitope Mapping: For novel ASK3 antibodies, employ tools like MAbTope to predict conformational epitopes .
Phosphorylation Studies: Use Phos-tag gels to resolve ASK3 activation states in osmotic stress assays .

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ASK3 Antibody