CSNK1G1 Antibody

What is CSNK1G1 and what cellular functions does it regulate?

CSNK1G1 (Casein kinase I isoform gamma-1) is a member of the protein kinase superfamily that functions as a serine/threonine kinase involved in multiple cellular signaling pathways. It regulates critical cellular processes including cell cycle progression, proliferation, and invasion. The circular RNA form (circ-CSNK1G1) has been found to be significantly overexpressed in thyroid cancer tissues compared to normal tissues, suggesting its role in cancer development and progression . Research indicates that CSNK1G1 participates in regulatory networks involving miRNAs, particularly through the miR-149-5p/MAPK1 pathway in cancer cells .

Which experimental applications are most suitable for CSNK1G1 antibodies?

CSNK1G1 antibodies have been validated for multiple experimental techniques including:

• Enzyme-Linked Immunosorbent Assay (ELISA): Useful for quantitative detection of CSNK1G1 expression levels in tissue or cell lysates

• Western Blotting (WB): Effective for detecting CSNK1G1 protein expression and analyzing post-translational modifications

• Immunohistochemistry (IHC): Valuable for visualizing CSNK1G1 expression patterns in tissue sections and determining subcellular localization

For phospho-specific antibodies like phospho-CSNK1G1 (Y263), immunohistochemistry has proven particularly effective for detecting activated forms of the protein in tissues such as brain samples .

How should CSNK1G1 antibodies be stored and handled to maintain reactivity?

CSNK1G1 antibodies are typically supplied in buffer solutions containing stabilizers like glycerol (often 50%), BSA (0.5%), and sodium azide (0.02%) . For optimal performance:

• Store antibodies at -20°C for long-term storage

• Avoid repeated freeze-thaw cycles by aliquoting the antibody

• Polyclonal antibodies like the phospho-CSNK1G1 (Y263) should be maintained in their liquid form in PBS-based buffers

• Always follow manufacturer-specific recommendations for storage conditions and shelf-life

When handling antibodies for experiments, maintain cold chain protocols and prepare working dilutions fresh for each experiment to ensure consistent results.

What are the optimal dilution ranges for CSNK1G1 antibodies in different applications?

Dilution requirements vary significantly depending on the application and specific antibody formulation:

Each new batch of antibody should undergo titration experiments to determine optimal working concentrations. For phospho-specific antibodies (e.g., phospho-Tyr263 CSNK1G1), lower dilutions may be required to detect phosphorylated epitopes effectively .

How can I validate the specificity of CSNK1G1 antibodies in my experimental system?

Multi-step validation is crucial for ensuring antibody specificity:

• Positive controls: Use tissues/cells known to express CSNK1G1 (e.g., brain tissue for phospho-Tyr263 CSNK1G1)

• Negative controls: Include samples where the protein is absent or knockdown models

• Peptide competition assays: Pre-incubation with the immunizing peptide should abolish specific signals

• Phosphatase treatment: For phospho-specific antibodies, treating samples with phosphatase should eliminate signal

• Cross-reactivity assessment: Test against related isoforms (CSNK1G2, CSNK1G3) if working with isoform-specific antibodies

The phospho-Tyr263 CSNK1G1 antibody specificity can be validated through peptide blocking experiments as demonstrated in immunohistochemistry analyses of human brain tissue .

What sample preparation methods maximize CSNK1G1 detection in tissue samples?

For optimal CSNK1G1 detection in tissue samples:

• Fixation: 10% neutral buffered formalin fixation for 24-48 hours depending on tissue size

• Antigen retrieval: Heat-induced epitope retrieval using citrate buffer (pH 6.0) or EDTA buffer (pH 9.0) is often effective

• Blocking: Use 5-10% normal serum from the same species as the secondary antibody

• Incubation: Overnight incubation at 4°C often yields better results than shorter incubations

• For phospho-specific detection: Include phosphatase inhibitors (e.g., sodium orthovanadate, sodium fluoride) in all buffers to preserve phosphorylation status

When working with paraffin-embedded samples, complete deparaffinization and adequate hydration are essential for antibody accessibility to the target epitope .

How can I analyze CSNK1G1's role in cancer progression using antibody-based approaches?

To investigate CSNK1G1's role in cancer progression:

• Expression analysis: Compare CSNK1G1 levels between tumor and matched normal tissues using IHC and Western blotting

• Functional studies: Combine CSNK1G1 knockdown/overexpression with antibody detection to correlate expression with phenotypic changes

• Phosphorylation status: Use phospho-specific antibodies to assess activation status in different tumor grades

• Co-immunoprecipitation: Identify CSNK1G1 interacting partners using antibody-based pull-down assays

• Chromatin immunoprecipitation (ChIP): If studying transcriptional effects associated with CSNK1G1 signaling

Research on thyroid cancer has demonstrated that circ-CSNK1G1 functions in the miR-149-5p/MAPK1 axis, promoting cancer cell proliferation, survival, and invasion . Similar approaches can be extended to other cancer types to elucidate CSNK1G1's role in disease mechanism.

What strategies address non-specific binding when using CSNK1G1 antibodies?

Non-specific binding can be minimized through:

• Titration optimization: Test multiple antibody concentrations to find the optimal signal-to-noise ratio

• Blocking enhancement: Extend blocking time or try alternative blocking agents (BSA, normal serum, commercial blockers)

• Washing optimization: Increase number and duration of washes using detergents like Tween-20

• Secondary antibody cross-reactivity: Pre-adsorb secondary antibodies or switch to highly cross-adsorbed alternatives

• Sample-specific considerations: For tissues with high endogenous peroxidase activity, use dual peroxidase blocking

If non-specific nuclear staining occurs in IHC, reducing primary antibody concentration and extending the blocking step often improves results. For phospho-specific antibodies like phospho-Tyr263 CSNK1G1, validation with phospho-peptide blocking is essential to distinguish specific from non-specific signals .

How do post-translational modifications affect CSNK1G1 antibody recognition?

Post-translational modifications (PTMs) significantly impact antibody-epitope interactions:

• Phosphorylation: Phospho-specific antibodies like anti-phospho-Tyr263 CSNK1G1 recognize only the phosphorylated form of the protein

• Conformational changes: PTMs may induce structural changes that mask or expose epitopes

• Antibody selection: For comprehensive protein analysis, use multiple antibodies targeting different regions of CSNK1G1

• Sample preparation: Preserving PTMs requires specific buffers with appropriate inhibitors

• Detection strategies: Consider using both phospho-specific and total protein antibodies to calculate activation ratios

For research focusing on CSNK1G1 activation status, phospho-Tyr263 antibodies provide critical information about kinase activity state that total protein antibodies cannot reveal .

What is the role of CSNK1G1 in thyroid cancer pathogenesis?

CSNK1G1, particularly its circular RNA form (circ-CSNK1G1), plays a significant role in thyroid cancer development:

• Expression pattern: Circ-CSNK1G1 is markedly upregulated in thyroid cancer tissues compared to normal tissues

• Molecular mechanism: Circ-CSNK1G1 functions as a miR-149-5p sponge, relieving miR-149-5p inhibition on MAPK1

• Cellular effects: Knockdown of circ-CSNK1G1 inhibits cancer cell proliferation, colony formation, survival, and invasion

• Cell cycle regulation: Circ-CSNK1G1 silencing arrests cells at the G0/G1 stage and reduces expression of cell cycle regulators like CyclinD1

• In vivo effects: Circ-CSNK1G1 knockdown suppresses tumor growth in mouse models

The mechanistic pathway involves circ-CSNK1G1 acting as a competing endogenous RNA that binds to miR-149-5p, preventing its inhibitory effect on MAPK1, thus promoting cancer progression .

How can CSNK1G1 antibodies be used to study its interaction with miRNAs and downstream targets?

To investigate CSNK1G1's interactions with miRNAs and downstream targets:

• RNA immunoprecipitation (RIP): Use CSNK1G1 antibodies to pull down associated RNA complexes, followed by qPCR to detect interacting miRNAs

• Dual detection: Combined fluorescent in situ hybridization (FISH) for miRNAs and immunofluorescence for CSNK1G1 protein

• Expression correlation: Analyze relationships between CSNK1G1, miR-149-5p, and MAPK1 expression using antibody-based detection methods

• Functional validation: Combine knockdown/overexpression studies with antibody detection of pathway components

• Immunohistochemistry multiplex: Sequential staining for CSNK1G1 and downstream targets like MAPK1 in the same tissue sections

Research has demonstrated a negative correlation between circ-CSNK1G1 and miR-149-5p expression in thyroid cancer tissues, with mechanistic studies confirming that circ-CSNK1G1 knockdown increases miR-149-5p levels and decreases MAPK1 expression .

What experimental approaches can differentiate between CSNK1G1 isoforms in tissue samples?

Distinguishing between CSNK1G1 isoforms requires strategic experimental design:

• Isoform-specific antibodies: Select antibodies targeting unique regions not conserved between isoforms

• Western blot analysis: Isoforms with different molecular weights can be distinguished by their migration patterns

• RT-PCR analysis: Design primers targeting isoform-specific exons to complement protein-level analysis

• Bioinformatic support: Use sequence alignment tools to identify unique epitopes for antibody selection

• Knockout/knockdown validation: Use isoform-specific silencing to confirm antibody specificity

For distinguishing between CSNK1G1, CSNK1G2, and CSNK1G3, careful antibody selection is critical. Some antibodies like phospho-Tyr263 may recognize conserved regions across multiple isoforms, while others may be truly isoform-specific .

How might new CSNK1G1 antibody technologies enhance cancer biomarker development?

Emerging antibody technologies offer promising avenues for CSNK1G1-based cancer biomarker development:

• Single-cell antibody-based proteomics: Analyzing CSNK1G1 expression at single-cell resolution to identify tumor heterogeneity

• Multiplexed immunofluorescence: Simultaneous detection of CSNK1G1 and multiple pathway components to create signature profiles

• Machine learning integration: Combining antibody-based tissue imaging with AI algorithms for automated pattern recognition

• Liquid biopsy applications: Developing highly sensitive antibody-based assays for detecting CSNK1G1 in circulating tumor cells

• Spatial transcriptomics correlation: Integrating antibody-based protein detection with spatial RNA analysis

Given circ-CSNK1G1's established role in thyroid cancer , these technologies could help develop diagnostic and prognostic tools based on CSNK1G1 pathway alterations.

What methodological approaches could improve detection of circular RNA-encoded CSNK1G1 variants?

Detecting circular RNA-encoded CSNK1G1 variants requires specialized approaches:

• Junction-specific antibodies: Develop antibodies targeting novel epitopes created at circular RNA junction sites

• Combined RNA-protein detection: Implement RNAscope technology alongside antibody-based protein detection

• Circular RNA-specific protocols: Use RNase R digestion to enrich for circular RNAs prior to analysis

• Back-splice junction primers: Design PCR primers to specifically amplify circular RNA forms

• Mass spectrometry validation: Confirm antibody-detected proteins through peptide mapping

Studies on circ-CSNK1G1 in thyroid cancer demonstrate that circular RNA forms are resistant to RNase R digestion and lack poly(A) tails, distinguishing them from linear transcripts .

How can phosphorylation-specific CSNK1G1 antibodies illuminate kinase activation networks in cancer?

Phosphorylation-specific antibodies provide unique insights into CSNK1G1 activation states:

• Dynamic phosphorylation mapping: Track changes in CSNK1G1 phosphorylation in response to stimuli or drug treatments

• Phosphorylation-dependent interactions: Identify proteins that interact specifically with phosphorylated CSNK1G1

• Pathway activation signatures: Create activation profiles by simultaneously detecting multiple phosphorylated proteins

• Kinase inhibitor assessment: Evaluate efficacy of kinase inhibitors by monitoring phosphorylation status

• Intratumoral heterogeneity: Analyze spatial distribution of phosphorylated CSNK1G1 within tumor microenvironments

Phospho-specific antibodies like phospho-Tyr263 CSNK1G1 can reveal activation patterns in tissues that correlate with disease states, potentially guiding targeted therapy approaches .