The Application and Validation of Serine/Threonine Phosphorylation Antibodies in Signal Transduction Research

I. Introduction

Protein phosphorylation is one of the most widespread and important post-translational modifications in eukaryotic cells, playing a central role in regulating life processes such as cell proliferation, differentiation, metabolism, and apoptosis. Serine (Ser) and threonine (Thr) phosphorylation account for more than 90% of all phosphorylation events and are key regulatory mechanisms in various signaling pathways such as MAPK, PI3K/AKT, and mTOR. As research on phosphorylation functions deepens, high-quality, highly specific phospho-specific antibodies have become indispensable tools in signal transduction research, disease mechanism exploration, and drug target validation.

II. Biological Significance of Serine/Threonine Phosphorylation

Serine/threonine phosphorylation refers to the covalent attachment of a phosphate group to the side-chain hydroxyl group of serine or threonine residues. This process is catalyzed by Ser/Thr kinases and can be reversed by Ser/Thr phosphatases. This modification enables precise regulation of cellular functions by altering protein conformation, activity, stability, or subcellular localization.

Typical pathways include:

MAPK/ERK pathway: regulates cell growth and differentiation;

PI3K/AKT pathway: involved in cell survival and metabolism;

mTOR pathway: controls protein synthesis and autophagy;

CDK/cyclin complexes: drive cell cycle progression;

AMPK pathway: senses energy status and regulates metabolic balance.

III. Development and Validation of Serine/Threonine Phosphorylation Antibodies

1. Antibody Types
Start Bio offers various types of phosphorylation antibodies, including:
Pan-serine/threonine phosphorylation antibodies: recognize multiple phospho-Ser/Thr sites, suitable for preliminary screening;
Site-specific phosphorylation antibodies: target specific phosphorylation sites on specific proteins, such as Akt Ser473, GSK-3β Ser9, p44/42 MAPK Thr202/Tyr204, etc.

2. Validation Methods and Data
Taking the Phospho-Serine/Threonine Recombinant Mouse mAb (S0B6454) as an example, this antibody demonstrates good specificity and sensitivity in Western Blot experiments:

Experimental conditions: 5% NFDM/TBST was used as the blocking/dilution buffer, with a primary antibody dilution ratio of 1:1000;
Sample treatment: comparison of untreated and Calyculin A-treated HeLa cell lysates;
Results: multiple phosphorylated protein bands were observed in the Calyculin A-treated group, indicating that the antibody can broadly recognize Ser/Thr phosphorylated proteins.

3. Specificity and Cross-Species Reactivity
Most Start Bio phosphorylation antibodies have been validated in samples from humans, mice, rats, and other species. Some antibodies (e.g., Phospho-Akt Ser473) exhibit excellent specificity across various tissues.

IV. Application Cases in Research and Industry

1. Tumor Signaling Pathway Research
Akt signaling pathway: Phospho-Akt (Ser473) antibody is used to assess the activation status of the PI3K/AKT pathway, commonly seen in breast cancer, lung cancer, and other tumor models;
mTOR pathway: Combined use of Phospho-mTOR (Ser2448) and Phospho-4E-BP1 (Thr37/46) antibodies can reveal mTORC1 activity status.

2. Cell Cycle and Apoptosis Research
Histone H3 (Ser10) phosphorylation antibody is used to label mitotic cells;
Phospho-SAPK/JNK (Thr183/Tyr185) is used to study stress-induced apoptosis processes.

3. Neuroscience
Multi-site Tau protein phosphorylation antibodies can be used to study neurofibrillary tangles in Alzheimer's disease models;
CaMKII (Thr286) phosphorylation antibody is used to study synaptic plasticity.

4. Metabolic Research
AMPKα (Thr172) phosphorylation antibody is used to assess cellular energy status;
GSK-3β (Ser9) phosphorylation status is closely related to the insulin signaling pathway.

V. Product Advantages and Technical Support

Start Bio has developed a series of high-performance phosphorylation antibodies based on a recombinant rabbit monoclonal antibody platform, featuring:
High specificity: validated across multiple platforms such as IHC, WB, and IP;
High affinity: most antibodies achieve picomolar-level KD values;
Batch consistency: recombinant technology ensures stable performance across different batches;
Broad species compatibility: suitable for various experimental models including humans, mice, and rats.

VI. Start Bio Product Recommendations

Start Bio offers serine/threonine phosphorylation antibodies