LRPPRC Antibody, Biotin conjugated

What is LRPPRC and what is its biological significance?

LRPPRC (Leucine-rich pentatricopeptide repeat containing protein) is a 130 kDa mitochondrial protein that plays a crucial role in cellular function. It is primarily characterized as a cytoplasmic mRNA-binding protein involved in the processing of mitochondrial DNA transcripts . The protein contains distinctive leucine-rich pentatricopeptide repeat motifs and is expressed in various tissues throughout the body. LRPPRC is also known by several aliases including LRP130, GP130, and LSFC in scientific literature . Its significant involvement in mitochondrial function makes it a target of interest for researchers studying cellular metabolism and mitochondrial diseases.

What is the connection between LRPPRC mutations and human disease?

Mutations in the LRPPRC gene have been definitively linked to the French-Canadian type of Leigh syndrome, a severe neurological disorder characterized by lesions in the subcortical region of the brain . This connection highlights the critical nature of LRPPRC in normal neurological development and function. Research into LRPPRC has expanded our understanding of mitochondrial diseases, particularly those affecting energy metabolism in neural tissues. Investigating LRPPRC using specific antibodies provides researchers with tools to better understand the pathophysiological mechanisms underlying these conditions and potentially develop therapeutic approaches.

How does LRPPRC function at the molecular level?

At the molecular level, LRPPRC functions as an mRNA-binding protein that participates in the processing and stabilization of mitochondrial transcripts . Additionally, it has been demonstrated that LRPPRC interacts with the low-affinity receptor for leukemia inhibitory factor, triggering an intracellular signal cascade . This dual functionality positions LRPPRC at the intersection of mitochondrial gene expression regulation and cellular signaling pathways, making it a complex and important target for fundamental research in cellular biology and disease mechanisms.

What are the key specifications of commercially available LRPPRC Antibody, Biotin conjugated?

Commercial LRPPRC Antibody, Biotin conjugated products typically present the following specifications:

• Antibody Type: Polyclonal antibody

• Host Species: Primarily rabbit-derived

• Target Specificity: Human LRPPRC, with some products showing cross-reactivity with mouse and rat orthologs

• Molecular Weight: The target protein is approximately 157.9 kilodaltons

• Conjugation: Biotin-conjugated for enhanced detection capabilities

• Clonality: Polyclonal, offering recognition of multiple epitopes

• Purification Method: Typically protein A or G purified (>95% purity)

• Form: Supplied in liquid form with preservatives and stabilizers

These specifications provide researchers with critical information for experimental planning and antibody selection based on their specific research requirements.

What applications are supported by LRPPRC Antibody, Biotin conjugated?

LRPPRC Antibody, Biotin conjugated supports multiple experimental applications, with varying levels of validation across different suppliers. The primary applications include:

The biotin conjugation offers significant advantages in detection sensitivity across these applications, particularly for methods utilizing streptavidin-based detection systems .

What is the significance of the biotin conjugation for research applications?

The biotin conjugation of LRPPRC antibodies provides several methodological advantages:

• Enhanced Signal Amplification: The strong affinity between biotin and streptavidin (Kd ≈ 10^-15 M) enables significant signal amplification in detection systems.

• Versatile Detection Options: Biotin-conjugated antibodies can be paired with various streptavidin-conjugated detection reagents (HRP, fluorophores, gold particles), offering flexibility across experimental platforms.

• Reduced Background: In multi-step detection protocols, biotin-conjugated primary antibodies can eliminate the need for species-specific secondary antibodies, potentially reducing non-specific binding.

• Compatibility with Multiplexing: The biotin-streptavidin system can be incorporated into multiplexed detection strategies, allowing simultaneous analysis of multiple targets.

These advantages make biotin-conjugated LRPPRC antibodies particularly valuable for complex experimental designs requiring high sensitivity and specificity .

What is the optimal protocol for using LRPPRC Antibody, Biotin conjugated in ELISA?

For optimal ELISA results with LRPPRC Antibody, Biotin conjugated, the following protocol is recommended:

• Plate Preparation:

  • Coat high-binding 96-well plates with capture antigen (recombinant LRPPRC or sample lysate) at 1-10 μg/ml in carbonate buffer (pH 9.6)

  • Incubate overnight at 4°C

  • Wash 3× with PBS-T (PBS + 0.05% Tween-20)

• Blocking:

  • Block with 1-5% BSA in PBS for 1-2 hours at room temperature

  • Wash 3× with PBS-T

• Primary Antibody Incubation:

  • Apply LRPPRC Antibody, Biotin conjugated at 1:300-1:5000 dilution in blocking buffer

  • Incubate for 1-2 hours at room temperature or overnight at 4°C

  • Wash 5× with PBS-T

• Detection:

  • Add streptavidin-HRP at 1:1000-1:5000 dilution

  • Incubate for 30-60 minutes at room temperature

  • Wash 5× with PBS-T

  • Develop with appropriate substrate (TMB)

  • Stop reaction and read absorbance at 450 nm

• Optimization Parameters:

  • Antibody titration is essential to determine optimal concentration

  • Include both positive and negative controls

  • Consider using recombinant LRPPRC protein (901-1036AA region) as a standard

This protocol may require modification based on specific experimental conditions and equipment.

What are the critical considerations for Western blotting with LRPPRC Antibody, Biotin conjugated?

When performing Western blotting for LRPPRC detection, researchers should consider:

• Sample Preparation:

  • Lyse cells/tissues in RIPA buffer supplemented with protease inhibitors

  • For mitochondrial proteins, consider mitochondrial isolation prior to lysis

  • Heat samples at 95°C for 5 minutes in reducing sample buffer

• Gel Selection:

  • Use 8-10% SDS-PAGE gels to effectively resolve the large LRPPRC protein (157.9 kDa)

  • Consider gradient gels (4-15%) for improved resolution

• Transfer Conditions:

  • For large proteins like LRPPRC, use wet transfer at reduced voltage (30V) overnight at 4°C

  • Add 0.1% SDS to transfer buffer to facilitate movement of large proteins

• Blocking and Antibody Incubation:

  • Block membrane with 5% non-fat milk or 3% BSA in TBS-T

  • Dilute LRPPRC Antibody, Biotin conjugated at 1:300-1:5000 in blocking buffer

  • Incubate overnight at 4°C with gentle agitation

• Detection:

  • Incubate with streptavidin-HRP (1:5000-1:10000)

  • Develop using enhanced chemiluminescence

  • For quantitative analysis, consider exposure across multiple timepoints

• Controls and Validation:

  • Include positive control (tissue/cell line with high LRPPRC expression)

  • Consider LRPPRC knockdown or knockout samples as negative controls

  • Verify expected molecular weight (approximately 158 kDa)

Optimizing these parameters will enhance detection specificity and sensitivity.

How should immunohistochemistry protocols be modified for LRPPRC Antibody, Biotin conjugated?

For immunohistochemistry using LRPPRC Antibody, Biotin conjugated:

• Tissue Preparation:

  • Fix tissues in 10% neutral buffered formalin

  • Process and embed in paraffin

  • Section at 4-6 μm thickness

• Antigen Retrieval (critical for LRPPRC detection):

  • Heat-induced epitope retrieval in citrate buffer (pH 6.0) or EDTA buffer (pH 9.0)

  • Boil for 15-20 minutes, then cool gradually

• Endogenous Biotin Blocking (essential with biotin-conjugated antibodies):

  • Block endogenous biotin using an avidin/biotin blocking kit

  • This step is crucial to prevent false-positive signals

• Antibody Incubation:

  • Apply LRPPRC Antibody, Biotin conjugated at 1:200-1:400 dilution

  • Incubate overnight at 4°C in a humidified chamber

• Detection System:

  • Use streptavidin-HRP conjugate

  • Develop with DAB or other appropriate chromogen

  • Counterstain with hematoxylin

• Special Considerations:

  • Given LRPPRC's mitochondrial localization, expect predominantly cytoplasmic staining

  • Include known positive tissue controls (e.g., liver, kidney)

  • Consider dual-staining with mitochondrial markers for co-localization studies

Appropriate controls and careful optimization of the protocol will ensure reliable and reproducible results.

What are common technical challenges when working with LRPPRC Antibody, Biotin conjugated?

Researchers may encounter several challenges when working with LRPPRC Antibody, Biotin conjugated:

• High Background Signal:

  • Potential causes: Insufficient blocking, endogenous biotin interference, cross-reactivity

  • Solutions: Extend blocking time, implement avidin-biotin blocking, increase wash stringency

• Weak or Absent Signal:

  • Potential causes: Protein degradation, insufficient antigen retrieval, low expression levels

  • Solutions: Verify protein integrity via Ponceau staining, optimize antigen retrieval, increase antibody concentration, extend incubation time

• Multiple Bands in Western Blot:

  • Potential causes: Protein degradation, splice variants, cross-reactivity

  • Solutions: Use fresh samples with protease inhibitors, verify specificity against recombinant standards

• Storage-Related Issues:

  • Antibody activity may decrease with repeated freeze-thaw cycles

  • Solution: Aliquot antibody upon receipt and store at -20°C or -80°C as recommended

• Reproducibility Issues:

  • Potential causes: Lot-to-lot variability, protocol inconsistencies

  • Solutions: Maintain detailed protocol records, consider testing different lots side-by-side

Understanding these common challenges can help researchers develop effective troubleshooting strategies.

How can researchers validate the specificity of LRPPRC Antibody, Biotin conjugated?

Validation of LRPPRC Antibody, Biotin conjugated specificity is critical for reliable experimental results. Recommended validation approaches include:

• Positive and Negative Controls:

  • Test with known LRPPRC-expressing tissues/cells (positive control)

  • Compare with LRPPRC-knockdown or knockout samples (negative control)

• Peptide Competition Assay:

  • Pre-incubate antibody with immunizing peptide (e.g., recombinant human LRPPRC protein fragment 901-1036AA)

  • Observe elimination of specific signal

• Multiple Antibody Comparison:

  • Compare results with other validated LRPPRC antibodies targeting different epitopes

  • Consistent detection patterns support specificity

• Orthogonal Method Verification:

  • Confirm protein expression using RNA-based methods (qPCR, RNA-seq)

  • Verify subcellular localization using fractionation and complementary techniques

• Cross-Species Reactivity Assessment:

  • Test reactivity in recommended species (human, mouse, rat)

  • Evaluate alignment of antibody epitope sequence across species

• Mass Spectrometry Confirmation:

  • For ultimate validation, perform immunoprecipitation followed by mass spectrometry

These validation approaches ensure experimental results accurately reflect LRPPRC biology rather than technical artifacts.

How should researchers interpret variations in LRPPRC detection patterns across different tissues?

When interpreting variations in LRPPRC detection across tissues:

• Expression Level Differences:

  • LRPPRC expression varies naturally between tissues

  • Higher expression typically observed in metabolically active tissues (heart, liver, kidney)

  • Quantify relative expression using appropriate controls and normalization

• Localization Pattern Variations:

  • Predominantly mitochondrial localization expected

  • Some nuclear localization may be observed in certain cell types

  • Cytoplasmic vs. nuclear distribution may reflect functional states

• Post-translational Modifications:

  • Variations in apparent molecular weight may indicate tissue-specific post-translational modifications

  • Consider phosphorylation or other modifications that may alter antibody recognition

• Interaction Partners:

  • Co-immunoprecipitation results may reveal tissue-specific interaction partners

  • These differences can provide insights into tissue-specific functions of LRPPRC

• Disease-Associated Changes:

  • Compare normal vs. pathological tissues carefully

  • Document both quantity and localization changes

  • Consider relationship to mitochondrial dysfunction phenotypes

Careful consideration of these factors helps researchers differentiate between biological variations and technical artifacts when interpreting LRPPRC detection patterns.

How can LRPPRC Antibody, Biotin conjugated be utilized in co-immunoprecipitation studies?

Co-immunoprecipitation (Co-IP) with LRPPRC Antibody, Biotin conjugated offers powerful approaches to study protein-protein interactions:

• Standard Protocol:

  • Lyse cells in non-denaturing buffer (e.g., NP-40 buffer with protease inhibitors)

  • Pre-clear lysate with streptavidin beads

  • Incubate lysate with LRPPRC Antibody, Biotin conjugated overnight at 4°C

  • Add streptavidin beads and incubate 2-4 hours

  • Wash extensively and elute

  • Analyze by Western blot for potential interaction partners

• Advantages of Biotin-Conjugated Antibody for Co-IP:

  • Direct capture on streptavidin beads eliminates need for protein A/G

  • Reduced background from heavy and light chains in western blot analysis

  • Strong biotin-streptavidin interaction enhances pull-down efficiency

• Recommended Controls:

  • Input control (5-10% of lysate)

  • IgG-biotin control (non-specific rabbit IgG)

  • Reverse Co-IP with antibodies against suspected interaction partners

• Potential Interaction Partners to Investigate:

  • Mitochondrial RNA processing machinery components

  • Translation factors

  • Leukemia inhibitory factor receptor (documented interaction)

  • Other mitochondrial proteins implicated in Leigh Syndrome

This approach can reveal novel insights into LRPPRC's role in mitochondrial function and disease mechanisms.

What approaches can be used to study LRPPRC in mitochondrial RNA processing?

To investigate LRPPRC's role in mitochondrial RNA processing:

• RNA Immunoprecipitation (RIP):

  • Use LRPPRC Antibody, Biotin conjugated to immunoprecipitate LRPPRC-RNA complexes

  • Extract RNA from immunoprecipitates

  • Analyze bound RNAs through RT-qPCR or RNA sequencing

  • Compare mitochondrial transcript profiles in normal vs. disease conditions

• Mitochondrial Isolation Combined with Antibody Detection:

  • Isolate intact mitochondria using differential centrifugation

  • Perform submitochondrial fractionation

  • Detect LRPPRC localization within mitochondrial compartments

  • Correlate with RNA distribution and processing intermediates

• Proximity Labeling Approaches:

  • Generate LRPPRC fusion with biotin ligase (BioID)

  • Identify proteins in close proximity to LRPPRC in living cells

  • Validate interactions with co-immunoprecipitation using LRPPRC Antibody, Biotin conjugated

• LRPPRC Knockdown/Knockout Studies:

  • Deplete LRPPRC using siRNA or CRISPR/Cas9

  • Analyze consequences on mitochondrial transcript stability and processing

  • Use LRPPRC Antibody, Biotin conjugated to confirm knockdown efficiency

These approaches can provide mechanistic insights into LRPPRC's function in mitochondrial RNA metabolism.

How can researchers use LRPPRC Antibody, Biotin conjugated to investigate Leigh Syndrome pathophysiology?

For investigating Leigh Syndrome pathophysiology using LRPPRC Antibody, Biotin conjugated:

• Patient-Derived Cell Studies:

  • Compare LRPPRC expression, localization, and interaction partners in patient vs. control cells

  • Correlate alterations with mitochondrial dysfunction markers

  • Use LRPPRC Antibody, Biotin conjugated for both detection and pull-down applications

• Animal Model Analysis:

  • Examine LRPPRC expression in mouse models of mitochondrial disease

  • Perform immunohistochemistry of brain sections focusing on regions affected in Leigh Syndrome

  • Correlate LRPPRC levels with neurodegeneration markers

• Therapeutic Screening Applications:

  • Use LRPPRC detection as a biomarker in therapeutic compound screening

  • Monitor changes in LRPPRC localization, stability, or interaction partners

  • High-content imaging with fluorescent streptavidin detection of biotin-conjugated antibody

• Structure-Function Correlations:

  • Compare wild-type vs. mutant LRPPRC detection patterns

  • Investigate if mutations alter protein-protein interactions

  • Examine consequences on downstream mitochondrial functions

• Multi-omics Integration:

  • Correlate LRPPRC immunoprecipitation data with:

    • Transcriptomics (RNA-seq)

    • Proteomics (mass spectrometry)

    • Metabolomics (mitochondrial metabolites)

  • Build integrated models of LRPPRC function in health and disease

These approaches leverage the specificity and versatility of LRPPRC Antibody, Biotin conjugated to advance understanding of Leigh Syndrome pathophysiology.

What are emerging applications for LRPPRC Antibody, Biotin conjugated in research?

Emerging applications for LRPPRC Antibody, Biotin conjugated include:

• Single-Cell Analysis:

  • Combining with mass cytometry (CyTOF) for single-cell profiling

  • Integration with spatial transcriptomics to correlate LRPPRC protein levels with transcript distributions

• Super-Resolution Microscopy:

  • Visualizing LRPPRC distribution within mitochondrial subcompartments

  • Studying co-localization with mitochondrial RNA granules at nanoscale resolution

• In vivo Imaging:

  • Development of LRPPRC imaging probes based on antibody fragments

  • Monitoring mitochondrial stress responses in living systems

• Liquid Biopsy Development:

  • Exploring LRPPRC as a potential biomarker in circulating extracellular vesicles

  • Correlation with mitochondrial disease progression

• Therapeutic Target Validation:

  • Using LRPPRC antibodies to validate drug target engagement

  • Monitoring changes in LRPPRC-dependent pathways during experimental therapeutics

These emerging applications represent the frontier of LRPPRC research, where biotin-conjugated antibodies offer particular advantages due to their detection sensitivity and versatility.

What resources are available to researchers working with LRPPRC Antibody, Biotin conjugated?

Researchers working with LRPPRC Antibody, Biotin conjugated can access:

• Commercial Resources:

  • Multiple vendors offering LRPPRC antibodies with various conjugates

  • Recombinant LRPPRC proteins for positive controls

  • Validation data available from manufacturers

• Research Tools:

  • Cell lines with verified LRPPRC expression profiles

  • LRPPRC knockout/knockdown models

  • Sequence information and epitope mapping data

• Literature Resources:

  • Published protocols for LRPPRC detection in various applications

  • Comparative studies of different antibody clones

  • Disease-specific studies utilizing LRPPRC antibodies

• Bioinformatic Tools:

  • Protein interaction databases with LRPPRC networks

  • Mitochondrial proteome databases

  • Expression atlases across tissues and conditions

• Research Community:

  • Mitochondrial research consortia

  • Leigh Syndrome research networks

  • Method-sharing platforms and protocol repositories

Leveraging these resources can accelerate research progress and enhance experimental reproducibility in LRPPRC studies.