LRP11 Antibody

What is LRP11 and what are its primary biological functions?

LRP11 (Low-density lipoprotein receptor-related protein 11) is a member of the LDL receptor family essential for cholesterol homeostasis. Recent research has identified its significant roles in multiple cellular processes including immune response modulation and cancer progression. Specifically, LRP11 has been shown to interact with LDL and activate downstream signaling pathways, particularly the MAPK13-TCF1 pathway that enhances antitumor immunity through promoting stem-like T cells . In cancer research, LRP11 has emerged as a regulator of PD-L1 expression in prostate cancer through β-catenin activation and influences cell proliferation, migration, and invasion in cervical cancer . The multifunctional nature of LRP11 makes it a promising target for therapeutic development across several disease contexts.

What types of LRP11 antibodies are available for research applications?

Multiple types of LRP11 antibodies are commercially available for research applications. These include monoclonal and polyclonal antibodies optimized for various experimental techniques. According to the search results, Santa Cruz Biotechnology offers several options including antibodies suitable for Western blotting (under non-reducing conditions), immunofluorescence, immunohistochemistry, and flow cytometry . R&D Systems also produces antibodies used in immunohistochemical staining at concentrations of approximately 10 μg/mL . When selecting an LRP11 antibody, researchers should consider the specific application, species reactivity (human, mouse, rat), and the epitope recognition. The choice between monoclonal antibodies (offering high specificity but potentially limited epitope recognition) and polyclonal antibodies (with broader epitope recognition but potential batch variability) should be determined by your experimental design and validation requirements.

How should LRP11 antibodies be validated before experimental use?

Validation of LRP11 antibodies requires a multi-step approach to ensure specificity, sensitivity, and reproducibility:

• Specificity validation:

  • Western blot analysis comparing wild-type vs. LRP11 knockdown or knockout cells

  • Peptide competition assays to confirm epitope specificity

  • Testing multiple antibodies targeting different epitopes to confirm consistent results

• Sensitivity assessment:

  • Titration experiments to determine optimal antibody concentration

  • Testing across a range of known expression levels

• Cross-reactivity examination:

  • Testing in multiple species if cross-reactivity is claimed

  • Evaluating potential cross-reactivity with other LRP family members

• Application-specific validation:

  • For IHC: Include positive and negative control tissues

  • For IF: Include subcellular localization controls and co-staining with organelle markers

  • For FCM: Include proper gating controls and isotype controls

Research from multiple groups should be compared when possible, and consistent detection across different experimental conditions strengthens validation. Published literature using specific antibody catalog numbers (like sc-514698 from Santa Cruz) can provide precedent for successful applications .

How does LRP11 contribute to tumor immune evasion mechanisms?

LRP11 plays a dual role in tumor immunity, with context-dependent effects that can either enhance or suppress anti-tumor immune responses. In prostate cancer, LRP11 activates β-catenin signaling, which subsequently induces PD-L1 expression . This upregulation of PD-L1 can contribute to immune evasion by inhibiting T cell function through the PD-1/PD-L1 axis. In experimental prostate cancer models, LRP11 overexpression induced immunosuppression in co-culture systems with immune cells, an effect that could be blocked by neutralizing antibodies against either LRP11 or PD-L1 .

Conversely, in the tumor microenvironment, LRP11 activation in T cells can enhance anti-tumor immunity by promoting stem-like TCF1+PD1+CD8+ T cells through the MAPK13-TCF1 pathway . These stem-like T cells have enhanced capacity for self-renewal and sustained anti-tumor activity. Understanding this context-dependent regulation is crucial for developing targeted therapeutic approaches that can modulate LRP11 activity specifically in either tumor cells or immune cells.

What methodological approaches are effective for studying LRP11 in cancer models?

Several methodological approaches have been established to study LRP11 in cancer models:

• Expression analysis:

  • Immunohistochemical staining of tissue microarrays (TMAs) using validated LRP11 antibodies (10 μg/mL, R&D Systems, AF8355)

  • Comparative analysis between tumor and normal tissues

  • Correlation with other biomarkers (e.g., P16 in cervical cancer)

• Functional studies through gene manipulation:

  • RNA interference using LRP11-specific shRNA delivered via lentiviral vectors

  • CRISPR/Cas9 knockout systems using plasmids targeting LRP11

  • Gene activation systems using CRISPR activation plasmids and lentiviral particles

• Phenotypic assays following LRP11 modulation:

  • Cell proliferation (CCK-8 assays)

  • Migration and invasion assays

  • In vivo tumor growth models

• Mechanistic studies:

  • Pathway analysis focusing on β-catenin signaling

  • Immunomodulatory effects using co-culture systems with immune cells

  • Protein interaction studies to identify binding partners

The combined use of these approaches allows for comprehensive characterization of LRP11's role in cancer initiation, progression, and potential therapeutic targeting.

How can researchers effectively interpret LRP11 expression patterns in different cancer types?

Interpreting LRP11 expression patterns across cancer types requires careful consideration of multiple factors:

• Tissue-specific baselines:

  • Compare tumor LRP11 expression with matched normal tissues from the same organ

  • Consider cell type-specific expression patterns within heterogeneous tissues

  • Establish threshold criteria that account for normal variation

• Correlation with clinical parameters:

  • Analyze association with stage, grade, and patient outcomes

  • Consider multivariate analysis to account for confounding factors

  • Evaluate correlation with established biomarkers (e.g., PD-L1 in prostate cancer )

• Technical considerations:

  • Use standardized scoring systems for immunohistochemistry

  • Consider different isoforms or post-translational modifications

  • Validate findings with multiple detection methods (protein vs. mRNA)

• Functional context:

  • Evaluate expression in relation to relevant signaling pathways (β-catenin, MAPK13-TCF1 )

  • Consider tumor microenvironment influences (immune infiltration, LDL availability)

  • Determine subcellular localization, which may indicate activation status

Rigorous statistical analysis and validation across independent cohorts are essential for establishing reliable LRP11 expression patterns as potential biomarkers or therapeutic targets in specific cancer contexts.

How does the LRP11-MAPK13-TCF1 signaling axis regulate T cell function?

The LRP11-MAPK13-TCF1 signaling axis represents a newly identified pathway that regulates T cell stemness and function in the context of tumor immunity. This pathway begins with LRP11 activation, typically through interaction with LDL, which subsequently induces MAPK13 activation . Once activated, MAPK13 is transported into the nucleus where it functions as a kinase, phosphorylating the transcription factor TCF1.

The significance of this pathway lies in its ability to promote the development and maintenance of stem-like T cells (TCF1+PD1+CD8+ T cells) within the tumor microenvironment. These stem-like T cells are particularly important for sustained anti-tumor immunity and have been identified as the T cell population that responds most effectively to checkpoint blockade immunotherapy .

In experimental settings, activation of this pathway enhances PD1 blockade immunotherapy, suggesting its potential for improving clinical outcomes in cancer treatment . The LRP11-MAPK13-TCF1 axis represents a novel target for immunomodulatory strategies aimed at enhancing T cell function in cancer.

What experimental approaches are used to analyze LRP11's impact on T cell stemness?

Analyzing LRP11's impact on T cell stemness requires a multi-faceted experimental approach:

• Phenotypic characterization:

  • Flow cytometry to quantify TCF1+PD1+CD8+ T cell populations following LRP11 activation or inhibition

  • Analysis of stemness markers (Slamf6, Bcl-2, IL-7R) and effector markers (Granzyme B, IFN-γ)

  • Serial adoptive transfer experiments to assess self-renewal capacity

• Signaling pathway analysis:

  • Phospho-flow or Western blotting to track MAPK13 activation and TCF1 phosphorylation

  • Nuclear/cytoplasmic fractionation to assess MAPK13 nuclear translocation

  • Chromatin immunoprecipitation to identify TCF1 binding sites after LRP11 activation

• Functional assessment:

  • In vitro proliferation assays using CD3/CD28 stimulation in the presence of LDL (80 μg/mL) and/or LRP11 antibodies (3 μg/mL)

  • Ex vivo re-stimulation assays to assess recall responses

  • In vivo tumor models to evaluate persistence and anti-tumor activity

• Transcriptional profiling:

  • RNA sequencing to assess gene expression changes following LRP11 modulation

  • Single-cell RNA-seq to characterize heterogeneity within T cell populations

  • ATAC-seq to identify changes in chromatin accessibility associated with stemness

These approaches provide complementary data on how LRP11 regulates the molecular and functional aspects of T cell stemness.

How do LRP11 agonists and antagonists differentially affect immune responses?

LRP11 agonists and antagonists exhibit distinct immunomodulatory effects that can be leveraged for different therapeutic contexts:

The context-dependent effects highlight the importance of targeted delivery and cell type-specific modulation when developing LRP11-targeting therapeutic strategies. Researchers should carefully consider the specific immune compartment they wish to modulate and select appropriate agonistic or antagonistic approaches accordingly.

How can researchers effectively combine LRP11 antibodies with genetic tools for comprehensive studies?

Researchers can maximize the utility of LRP11 antibodies by integrating them with complementary genetic tools in well-designed experimental approaches:

• Combining antibody-based detection with genetic manipulation:

  • Use LRP11 antibodies to confirm knockdown/knockout efficiency after employing shRNA (sc-40101-SH, sc-40102-SH) or CRISPR/Cas9-based approaches (sc-400638, sc-421464)

  • Validate antibody specificity using genetic knockdown/knockout controls

  • Combine overexpression studies with antibody-based detection to track protein localization changes

• Multi-modal analysis approaches:

  • Correlate immunohistochemical staining with transcript levels from the same samples

  • Use antibodies in flow cytometry to isolate LRP11-expressing cells for subsequent molecular analysis

  • Combine with proximity ligation assays to identify protein-protein interactions in situ

• Genetic rescue experiments:

  • Reintroduce wild-type or mutant LRP11 after knockdown/knockout to establish causality

  • Use domain-specific mutations to map functional regions required for specific signaling outcomes

  • Create chimeric receptors to identify domains required for LDL interaction and signal transduction

• Temporal control strategies:

  • Implement inducible expression/knockdown systems alongside antibody blockade

  • Compare acute (antibody-mediated) versus chronic (genetic) manipulation

  • Use optogenetic or chemogenetic approaches for precise temporal control of LRP11 activation

What advanced techniques can be used to study the dynamics of LRP11-mediated signaling?

Several advanced techniques can be employed to dissect the dynamics of LRP11-mediated signaling pathways:

• Phosphoproteomics analysis:

  • Mass spectrometry-based identification of phosphorylation changes following LRP11 activation or inhibition

  • Targeted phospho-specific Western blotting focusing on MAPK13 and TCF1 phosphorylation sites

• Chromatin immunoprecipitation sequencing (ChIP-seq):

  • Mapping TCF1 binding sites before and after LRP11 activation

  • Identifying changes in chromatin accessibility associated with LRP11-mediated transcriptional reprogramming

• Live-cell imaging techniques:

  • FRET-based biosensors to monitor real-time activation of MAPK13 following LRP11 engagement

  • Tracking nuclear translocation of signaling components using fluorescent fusion proteins

• Single-cell analysis approaches:

  • Single-cell RNA-seq to identify transcriptional changes in heterogeneous populations following LRP11 activation

  • Mass cytometry (CyTOF) to simultaneously examine multiple pathway components at the single-cell level

• In vivo pathway visualization:

  • Intravital microscopy combined with reporter systems to visualize pathway activation in tumor microenvironments

  • Genetic reporter mice expressing fluorescent proteins under the control of TCF1-responsive elements

These advanced techniques can provide deeper insights into the spatiotemporal dynamics and molecular mechanisms of LRP11-mediated signaling across different biological contexts.

How can researchers quantitatively assess LRP11 activation and signaling output?

Quantitative assessment of LRP11 activation requires multi-parametric approaches that capture both direct receptor engagement and downstream signaling events:

• Direct LRP11 activation metrics:

  • Receptor internalization assays using fluorescently-labeled LDL or antibodies

  • Conformational change detection using conformation-specific antibodies

  • FRET-based assays to detect LRP11 dimerization or clustering

• Downstream signaling readouts:

  • Quantitative Western blotting for phosphorylated MAPK13 with densitometric analysis

  • Nuclear/cytoplasmic fractionation followed by MAPK13 quantification to assess nuclear translocation

  • Quantitative PCR for known transcriptional targets of TCF1 after phosphorylation

• Functional assessment approaches:

  • TCF1+ T cell quantification via flow cytometry as a readout of pathway activation in immune contexts

  • Luciferase reporter assays using TCF1-responsive elements

  • PD-L1 expression quantification in cancer cells as a readout of pathway activation in certain cancer types

For each quantitative approach, researchers should establish dose-response relationships, kinetic profiles, and appropriate statistical analyses to determine significant changes in activation status. Standardization using positive controls (e.g., direct MAPK13 activators) helps normalize between experiments and across research groups.

What are common challenges with LRP11 antibodies and how can they be addressed?

Researchers may encounter several challenges when working with LRP11 antibodies:

• Specificity issues:

  • Challenge: Cross-reactivity with other LRP family members due to structural similarities

  • Solution: Validate antibodies using LRP11 knockdown/knockout samples as negative controls

  • Solution: Use multiple antibodies targeting different epitopes to confirm findings

• Detection sensitivity:

  • Challenge: Low expression levels in certain cell types or tissues

  • Solution: Optimize antibody concentration through titration experiments

  • Solution: Consider signal amplification methods (TSA for IHC/IF, enhanced chemiluminescence for WB)

• Epitope accessibility:

  • Challenge: Epitope masking due to protein-protein interactions or conformational changes

  • Solution: Test multiple fixation and antigen retrieval methods for IHC/IF

  • Solution: Compare reducing vs. non-reducing conditions for Western blotting

• Batch-to-batch variability:

  • Challenge: Inconsistent results between antibody lots

  • Solution: Validate each new lot against previous lots using known positive samples

  • Solution: Consider creating a standard curve with recombinant protein for quantitative applications

Addressing these challenges requires systematic troubleshooting and maintaining detailed records of experimental conditions and antibody performance.

How should researchers interpret conflicting data regarding LRP11 function?

When faced with conflicting data regarding LRP11 function, researchers should adopt a systematic approach to interpretation:

• Context-dependent analysis:

  • Recognize that LRP11 may have different functions in different cellular contexts (immune cells vs. cancer cells)

  • Examine tissue specificity, cell-type specificity, and species differences that might explain discrepancies

  • Consider microenvironmental factors that might influence LRP11 signaling (e.g., LDL availability)

• Methodological evaluation:

  • Compare experimental approaches used in conflicting studies (in vitro vs. in vivo, knockdown vs. antibody blocking)

  • Evaluate the specificity of tools used (validate antibody specificity, assess off-target effects of genetic approaches)

  • Consider differences in readout systems and their sensitivity/specificity

• Integration with existing knowledge:

  • Place conflicting findings in the context of known LRP family biology

  • Consider whether discrepancies reflect different aspects of a complex biological system

  • Examine temporal aspects of signaling that might explain apparently contradictory results

• Validation strategies:

  • Design experiments that directly address contradictions using multiple complementary approaches

  • Use genetic rescue experiments to confirm specificity of observed phenotypes

  • Consider collaboration with groups reporting conflicting data to standardize approaches

By systematically evaluating conflicting data, researchers can develop more nuanced understanding of LRP11 biology and its context-dependent functions.

What essential controls should be included in LRP11 antibody experiments?

When designing experiments involving LRP11 antibodies, several essential controls should be included:

• Specificity controls:

  • Positive control: Samples with confirmed LRP11 expression (cell lines, tissues)

  • Negative control: LRP11 knockdown or knockout samples generated using validated shRNA or CRISPR systems

  • Isotype control: Matched isotype antibody at equivalent concentration to rule out non-specific binding

  • Peptide competition: Pre-incubation with blocking peptide to confirm epitope specificity

• Technical controls:

  • Loading control: Housekeeping protein detection for Western blotting

  • Staining control: Counterstain to provide anatomical context in IHC/IF

  • Secondary-only control: Omission of primary antibody to assess secondary antibody background

  • Non-permeabilized control: For distinguishing surface from intracellular staining in IF/flow cytometry

• Experimental manipulation controls:

  • Vehicle control: For studies using LDL, oxidized LDL, or HDL treatments

  • Concentration gradient: Titration of antibody or ligand to establish dose-response

  • Time course: Multiple timepoints to capture dynamic changes in signaling

  • Pathway controls: Established activators/inhibitors of the studied pathway (e.g., MAPK13 inhibitors)

Careful documentation of all control results is essential for proper interpretation and troubleshooting of experimental findings related to LRP11 function.