PIK3R6 Antibody, Biotin conjugated

What is PIK3R6 and what cellular functions does it regulate?

PIK3R6 (Phosphoinositide-3-Kinase, Regulatory Subunit 6) functions as a regulatory subunit of PI3Kγ with established tumor-promoting properties in several cancer types. Research has identified PIK3R6 as a critical component in cellular signaling pathways that regulate proliferation, migration, invasion, and angiogenesis. Recent studies demonstrate that PIK3R6 enhances angiogenesis in hepatocellular carcinoma (HCC) through activation of the STAT3 signaling pathway, accelerating malignant progression . Additionally, PIK3R6 has been implicated in clear cell renal cell carcinoma (CCRCC) development, where its upregulation correlates with poorer patient survival outcomes . The protein plays a multifaceted role in regulating the PI3K signaling network, which influences numerous cellular functions including cell cycle progression, apoptosis resistance, and metastatic capability.

What are the technical specifications of commercially available PIK3R6 antibodies with biotin conjugation?

Commercially available PIK3R6 antibodies with biotin conjugation present the following validated specifications:

The biotin conjugation enhances detection sensitivity, providing flexibility in experimental design through compatibility with various streptavidin-based detection systems .

How should researchers design immunodetection protocols using biotin-conjugated PIK3R6 antibodies?

When designing immunodetection protocols with biotin-conjugated PIK3R6 antibodies, researchers should implement the following methodological approach:

For ELISA applications:

• Sample preparation: Process clinical specimens or cell lysates using standardized protocols to ensure protein integrity.

• Blocking optimization: Use appropriate blocking buffers (typically PBS with 1-5% BSA) to minimize non-specific binding.

• Antibody titration: Determine optimal concentration through serial dilutions (typically starting at 1:500-1:2000), as specified manufacturer guidance indicates optimal dilutions should be determined by the end user .

• Detection system: Employ streptavidin-HRP or other streptavidin-conjugated detection systems that leverage the high-affinity biotin-streptavidin interaction.

• Validation controls: Include positive controls (samples known to express PIK3R6), negative controls (samples known to lack PIK3R6), and technical controls (omitting primary or secondary reagents).

For tissue-based detection:
Researchers investigating PIK3R6 expression in cancer tissues should follow the methodology demonstrated in recent CCRCC studies, where immunohistochemistry successfully detected differential PIK3R6 expression between tumor and adjacent normal tissues .

What validation approaches confirm specificity of PIK3R6 detection in experimental systems?

Rigorous validation of PIK3R6 detection requires multi-modal confirmation approaches:

• RNA interference validation: Employ siRNA-mediated knockdown of PIK3R6 (si-PIK3R6) as demonstrated in recent CCRCC research. This approach provides functional validation by confirming antibody signal reduction corresponds with decreased PIK3R6 expression. In published studies, researchers validated transfection efficiency using both quantitative real-time PCR and western blot analysis, revealing significant reduction in both mRNA and protein levels of PIK3R6 following siRNA transfection .

• Complementary detection methods: Validate antibody-based findings using independent techniques:

  • qRT-PCR for mRNA expression

  • Western blotting for protein expression

  • Immunohistochemistry for tissue localization

• Positive and negative tissue controls: Include tissues with known PIK3R6 expression profiles. Recent studies identified significant elevation of PIK3R6 in CCRCC tissues compared to para-carcinoma normal kidney tissues, providing benchmark expression patterns .

How can PIK3R6 antibodies facilitate investigation of cancer progression mechanisms?

PIK3R6 antibodies serve as critical tools for elucidating cancer progression mechanisms through several advanced applications:

• Expression profiling across cancer stages: Recent CCRCC research demonstrated that heightened PIK3R6 expression significantly correlates with tumor stage, grade, subtype, and metastasis status . These findings suggest PIK3R6 antibodies can effectively discriminate disease progression patterns.

• Pathway interaction analysis: In HCC research, PIK3R6 was shown to enhance angiogenesis by activating the STAT3 signaling pathway . Researchers can employ biotin-conjugated PIK3R6 antibodies in co-immunoprecipitation studies to identify novel protein interaction partners within these signaling networks.

• Mechanistic loss-of-function studies: Following the methodology of recent CCRCC research, investigators can design experiments combining PIK3R6 knockdown with subsequent functional analysis using PIK3R6 antibodies to monitor expression changes. In published studies, si-PIK3R6 transfection markedly reduced cell viability in both 786-O and ACHN cell lines, with corresponding changes in PIK3R6 expression confirmed via antibody-based detection .

• Prognostic biomarker development: Multiple cohort analyses have established that high PIK3R6 expression in tumor tissues negatively correlates with patient prognosis . Biotin-conjugated PIK3R6 antibodies can facilitate development of standardized immunoassays for potential clinical application.

What role does PIK3R6 play in the STAT3 signaling pathway and angiogenesis?

PIK3R6 functions as a novel promoter of angiogenesis through its interaction with the STAT3 signaling pathway, particularly in hepatocellular carcinoma:

• Pathway activation: Research indicates PIK3R6 boosts STAT3 signaling pathway activity, resulting in persistent activation of STAT3 in HCC (compared to transient activation in normal physiological states) .

• Transcriptional regulation: Activated STAT3 promotes transcription of several genes, including VEGFA molecules, which are key mediators of angiogenesis .

• Disease progression impact: This PIK3R6-STAT3-VEGFA axis accelerates malignant progression in HCC, suggesting PIK3R6 as a potential therapeutic target .

To investigate these mechanisms, researchers should employ PIK3R6 antibodies in combination with:

• Phospho-STAT3 detection to assess activation status

• ChIP assays to evaluate STAT3 binding to target promoters

• Angiogenesis assays (tube formation, migration) to assess functional outcomes

• In vivo models to evaluate tumor vascularization

How should researchers address technical challenges when using biotin-conjugated PIK3R6 antibodies?

When encountering technical challenges with biotin-conjugated PIK3R6 antibodies, researchers should implement the following troubleshooting strategies:

• Signal specificity issues:

  • Validate antibody specificity through siRNA knockdown experiments as demonstrated in recent literature where si-PIK3R6#1 showed more pronounced inhibitory effects on PIK3R6 expression than si-PIK3R6#2 .

  • Optimize blocking conditions to reduce background (typically 1-5% BSA or serum from species unrelated to host).

  • Include isotype control antibodies to distinguish non-specific binding.

• Signal intensity problems:

  • Adjust antibody concentration through careful titration experiments.

  • Consider signal amplification methods compatible with biotin-streptavidin systems.

  • Ensure proper storage at -20°C with minimal freeze-thaw cycles to maintain antibody integrity .

• Cross-reactivity concerns:

  • Verify species reactivity, as commercially available PIK3R6 antibodies are specifically validated for human samples .

  • When working with non-human samples, conduct preliminary validation studies.

What methodological considerations are critical when analyzing PIK3R6 expression in patient samples?

Analysis of PIK3R6 expression in patient samples requires careful methodological consideration:

• Tissue handling and processing:

  • Standardize collection procedures to minimize pre-analytical variables.

  • Process matched tumor and adjacent normal tissues simultaneously to ensure comparable conditions.

• Quantification approaches:

  • Implement semi-quantitative scoring systems for immunohistochemistry as used in recent CCRCC studies that demonstrated significantly elevated PIK3R6 in tumor tissues compared to normal kidney tissues (p < .001) .

  • Consider digital image analysis for more objective quantification.

• Clinical correlation analysis:

  • Stratify patients by PIK3R6 expression levels to assess correlation with clinical parameters.

  • Perform multivariate analysis to control for confounding variables.

  • Design studies with adequate statistical power based on preliminary data from CCRCC research showing correlation between PIK3R6 expression and tumor stage, grade, and metastasis status .

How might PIK3R6 antibodies contribute to developing therapeutic approaches targeting PI3K signaling?

PIK3R6 antibodies can facilitate therapeutic development through:

• Target validation: Recent research demonstrates that PIK3R6 silencing reduces cell proliferation, migration, and invasion while inducing G0/G1 phase arrest and apoptosis in cancer cells . These findings position PIK3R6 as a potential therapeutic target.

• Mechanism elucidation: Antibody-based studies can further clarify how PIK3R6 activates the STAT3 signaling pathway in HCC, identifying potential intervention points .

• Biomarker development: PIK3R6 antibodies can help identify patient populations most likely to benefit from PI3K pathway inhibitors through expression profiling.

• Antibody-drug conjugate potential: The specificity of PIK3R6 antibodies could potentially be leveraged for targeted therapeutic delivery, though additional research is needed in this direction.

What emerging roles of PIK3R6 in different pathological conditions warrant further investigation?

Emerging evidence suggests PIK3R6 involvement in multiple pathological conditions beyond cancer:

• Preeclampsia: PIK3R6 has been implicated in preeclampsia-like conditions through its role in the PIK3R6/p-STAT3 signaling pathway .

• Cancer type-specific mechanisms: While PIK3R6's role is established in CCRCC and HCC, its function in other cancer types remains to be fully characterized, particularly regarding tissue-specific mechanisms .

• Resistance mechanisms: Potential involvement of PIK3R6 in resistance to current therapies targeting the PI3K pathway merits investigation.

• Non-STAT3 signaling interactions: Additional signaling nodes that interact with PIK3R6 could reveal new therapeutic vulnerabilities.

Each of these areas represents promising directions for research utilizing PIK3R6 antibodies to expand our understanding of this important regulatory protein and its therapeutic implications.