C10orf82 Antibody

What is C10orf82 and what antibody types are available for research?

C10orf82 (Chromosome 10 Open Reading Frame 82) is a human protein with a molecular weight of approximately 17.6 kDa. This protein, identified by UniProt code Q8WW14 and Gene ID 143379, remains largely uncharacterized in terms of functionality . Research into this protein primarily utilizes two major antibody types:

• Rabbit Polyclonal Antibodies: These recognize multiple epitopes across the C10orf82 protein, particularly the AA 1-154 region. They are typically raised in rabbits using recombinant human C10orf82 protein as the immunogen .

• Mouse Monoclonal Antibodies: Clone OTI1G4 is a well-characterized mouse monoclonal antibody with IgG1 isotype. This antibody is generated against full-length human recombinant C10orf82 protein and demonstrates high specificity for human C10orf82 .

The choice between these antibody types should be guided by experimental requirements, with polyclonal antibodies offering broader epitope recognition and monoclonal antibodies providing higher specificity.

What applications are validated for C10orf82 antibodies?

C10orf82 antibodies have been validated for multiple research applications:

The mouse monoclonal antibody is particularly well-suited for Western blot applications with a recommended dilution of 1:2000, while also performing effectively in immunofluorescence studies at 1:100 dilution . The rabbit polyclonal antibody has broader application potential including ELISA, Western blot, and immunohistochemistry . When designing experiments, researchers should consider these validated applications to ensure optimal results.

How should C10orf82 antibodies be properly stored and handled?

Proper storage and handling are crucial for maintaining antibody functionality and experimental reproducibility. C10orf82 antibodies require specific storage conditions:

• Storage Temperature: Store at -20°C to -80°C for long-term preservation . The mouse monoclonal antibody specifically requires cool storage conditions .

• Buffer Composition: Most C10orf82 antibodies are supplied in PBS (pH 7.3) containing preservatives and stabilizers. The rabbit polyclonal antibody typically contains 0.01 M PBS (pH 7.4), 0.03% Proclin-300, and 50% glycerol . The mouse monoclonal antibody is preserved in PBS with 1% BSA, 50% glycerol, and 0.02% sodium azide .

• Aliquoting Recommendation: To prevent protein degradation from repeated freeze-thaw cycles, aliquot the antibody into smaller volumes upon first thaw .

• Safety Considerations: Note that these antibodies contain preservatives (ProClin or sodium azide) that are hazardous substances requiring appropriate handling precautions by trained personnel .

Adherence to these storage and handling guidelines will maximize antibody shelf-life and ensure consistent experimental outcomes.

How can researchers validate the specificity of C10orf82 antibodies?

Validating antibody specificity is essential for generating reliable research data. For C10orf82 antibodies, several validation approaches are recommended:

• Knockout/Knockdown Controls: Compare antibody binding in wild-type samples versus those where C10orf82 expression has been eliminated or reduced through CRISPR-Cas9 knockout or siRNA knockdown.

• Recombinant Protein Controls: Use purified recombinant C10orf82 protein as a positive control in Western blot applications to confirm correct target molecular weight (17.6 kDa) .

• Peptide Competition Assay: Pre-incubate the antibody with excess purified C10orf82 protein (particularly the 1-154AA region) to block specific binding sites before application to samples . Reduction in signal indicates specific binding.

• Cross-Reactivity Assessment: Test antibody reactivity against related proteins or in non-target species. The available C10orf82 antibodies show reactivity to human and, in some cases, mouse samples .

• Multiple Antibody Approach: Compare results using both monoclonal and polyclonal antibodies targeting different epitopes of C10orf82 to confirm specificity of observed signals.

These validation procedures should be documented and reported in publications to enhance experimental reproducibility and data reliability.

What considerations should guide Western blot protocol optimization for C10orf82 detection?

Western blot optimization for C10orf82 detection requires attention to several key parameters:

• Sample Preparation:

  • Use appropriate lysis buffers containing protease inhibitors to prevent degradation of C10orf82

  • Recommended protein loading: 10-30 μg of total protein per lane

  • Include both reducing and non-reducing conditions in initial optimization to determine optimal detection

• Gel Selection and Transfer:

  • Use higher percentage gels (12-15%) for optimal resolution of the 17.6 kDa C10orf82 protein

  • Consider PVDF membranes for better protein retention of low molecular weight targets

  • Transfer at lower voltage for longer time to ensure complete transfer of small proteins

• Antibody Incubation:

  • Primary antibody dilutions: 1:2000 for both monoclonal and polyclonal antibodies

  • Incubation time: Overnight at 4°C for optimal signal-to-noise ratio

  • Include 5% non-fat dry milk or BSA in blocking and antibody dilution buffers

• Detection and Visualization:

  • Enhanced chemiluminescence (ECL) systems are suitable for detecting C10orf82

  • Extended exposure times may be necessary due to potentially low endogenous expression

• Controls:

  • Positive control: Cell lines known to express C10orf82

  • Negative control: Antibody diluent without primary antibody

  • Loading control: Housekeeping protein of different molecular weight than C10orf82

Following these guidelines will help researchers achieve specific and reproducible detection of C10orf82 protein via Western blotting.

How do experimental conditions affect C10orf82 antibody binding in immunofluorescence applications?

When using C10orf82 antibodies for immunofluorescence (IF) studies, several factors impact results:

• Fixation Method:

  • Paraformaldehyde (4%) fixation preserves epitope structure while maintaining cellular morphology

  • Methanol fixation may enhance detection of certain epitopes but can disrupt membrane structures

  • Comparative testing of fixation methods is recommended for novel cell types

• Permeabilization Optimization:

  • 0.1-0.3% Triton X-100 is generally effective for nuclear proteins

  • Gentler permeabilization with 0.1% saponin may be preferred for membrane-associated proteins

  • Duration and temperature of permeabilization affect antibody penetration

• Antibody Selection and Dilution:

  • The mouse monoclonal antibody (OTI1G4) is specifically validated for IF at 1:100 dilution

  • Consider using FITC-conjugated anti-C10orf82 antibodies for direct detection when available

• Signal Amplification:

  • For low-abundance proteins, consider tyramide signal amplification

  • Balance signal enhancement with potential background increase

• Controls and Counterstaining:

  • Include DAPI nuclear counterstain for localization reference

  • Validate subcellular localization with organelle markers if C10orf82 localization is being studied

Optimizing these parameters is essential for generating reliable and reproducible immunofluorescence data for C10orf82 localization studies.

How should researchers interpret inconsistent results with C10orf82 antibodies?

When faced with inconsistent results using C10orf82 antibodies, systematic troubleshooting is essential:

• Antibody Variability Analysis:

  • Compare lot-to-lot variation by testing multiple antibody batches

  • Document performance differences between polyclonal and monoclonal antibodies

  • Consider epitope accessibility variations in different sample types

• Experimental Condition Assessment:

  • Evaluate buffer compatibility with antibody performance

  • Test multiple blocking agents to reduce non-specific binding

  • Optimize incubation temperature and duration systematically

• Sample-Specific Considerations:

  • Check for post-translational modifications that may mask epitopes

  • Assess protein expression levels across different tissue/cell types

  • Verify sample integrity and storage conditions

• Common Sources of Discrepancy:

• Documentation Practices:

  • Maintain detailed records of all experimental variables

  • Document antibody performance across different applications

  • Consider publishing negative results to inform the research community

Systematic evaluation of these factors will help determine whether inconsistencies reflect biological reality or technical artifacts.

How can computational approaches enhance C10orf82 antibody specificity design?

Recent advances in computational biology offer promising approaches to enhance antibody specificity:

• Binding Mode Identification:

  • Computational models can identify different binding modes associated with specific ligands

  • Biophysics-informed modeling helps disentangle binding modes even for chemically similar epitopes

  • These models can predict outcomes for novel ligand combinations beyond experimental data

• Customized Specificity Profiles:

  • Energy function optimization can generate antibody sequences with predefined binding profiles

  • Cross-specific variants: Jointly minimize energy functions for desired ligands

  • Highly specific variants: Minimize energy for target ligand while maximizing for non-targets

• Implementation Methodology:

  • Train models using high-throughput sequencing data from phage display experiments

  • Apply biophysics principles to identify energy parameters associated with binding specificity

  • Validate computational predictions with targeted experimental testing

• Advantages over Traditional Methods:

  • Overcomes limitations of experimental selection methods in library size

  • Provides greater control over specificity profiles than conventional approaches

  • Enables generation of antibodies with novel binding properties not present in initial libraries

These computational approaches represent a significant advancement over traditional antibody development methods, especially for challenging targets like C10orf82 where high specificity is required.

What considerations should guide the design of multifunctional C10orf82 antibody experiments?

Researchers developing advanced applications with C10orf82 antibodies should consider several design elements:

• Antibody Conjugation Strategies:

  • Multiple conjugation options are available for C10orf82 antibodies, including FITC, HRP, and biotin

  • Each conjugation affects molecular properties and may alter binding characteristics

  • Validation of conjugated antibodies should be performed independently of unconjugated versions

• Multiplex Experimental Design:

  • When combining C10orf82 detection with other targets:

    • Select antibodies from different host species to avoid cross-reactivity

    • Verify spectral separation when using multiple fluorophores

    • Test for potential antibody interference through sequential staining controls

• Quantitative Analysis Approaches:

  • For precise quantification, consider:

    • Establishing standard curves with recombinant C10orf82 protein

    • Implementing digital image analysis for immunostaining quantification

    • Using internal references for normalization across experimental conditions

• Advanced Detection Methods:

  • Super-resolution microscopy techniques may reveal precise subcellular localization

  • Proximity ligation assays can identify protein-protein interactions involving C10orf82

  • FRET-based approaches can measure dynamic protein interactions in living cells

• Functional Characterization:

  • Antibody-mediated pulldown combined with mass spectrometry to identify interaction partners

  • ChIP-seq applications if C10orf82 has DNA-binding properties

  • Neutralization experiments to assess functional roles of specific protein domains

These considerations enable researchers to move beyond basic detection to functional characterization of C10orf82 in complex biological systems.

How can C10orf82 antibodies be integrated into advanced immunoassay platforms?

Integration of C10orf82 antibodies into advanced platforms requires systematic optimization:

• Flow Cytometry Applications:

  • The mouse monoclonal antibody (OTI1G4) has been validated for flow cytometry

  • Consider using directly conjugated antibodies to eliminate secondary antibody steps

  • Optimize fixation and permeabilization for intracellular detection

  • Include appropriate compensation controls when multiplexing with other markers

• High-Content Screening:

  • Leverage immunofluorescence capabilities for automated image-based screening

  • Develop robust quantification algorithms for C10orf82 expression or localization

  • Validate consistency across plate positions to minimize edge effects

  • Include positive and negative controls on each plate for normalization

• Antibody Arrays and Multiplex Assays:

  • Test for cross-reactivity with other antibodies in multiplex panels

  • Optimize antibody concentrations to ensure equivalent sensitivity across targets

  • Consider bead-based multiplexing systems for simultaneous detection of multiple targets

• Single-Cell Analysis Platforms:

  • Validate C10orf82 antibody performance at single-cell resolution

  • Establish protocols compatible with single-cell sequencing workflows

  • Benchmark against genomic data when assessing protein expression heterogeneity

• Emerging Technologies Integration:

  • Assess compatibility with microfluidic systems for automated processing

  • Evaluate performance in organ-on-chip models for functional studies

  • Test integration with CRISPR screening platforms for systematic functional analysis

Strategic validation across these platforms extends the utility of C10orf82 antibodies beyond conventional applications to cutting-edge research methodologies.

How might affinity engineering improve C10orf82 antibody performance?

Emerging antibody engineering approaches offer promising avenues for enhancing C10orf82 antibody functionality:

• Glycoengineering for Enhanced Functionality:

  • Afucosylated antibodies demonstrate increased binding to Fc-IgG receptor CD16A

  • This modification can enhance antibody-dependent cellular functions by up to 50-fold

  • Engineering glycosylation patterns can create antibodies with customized effector functions

• Fragment-Based Optimization:

  • Fab fragments may provide superior access to sterically hindered epitopes

  • Single-domain antibodies could offer increased tissue penetration

  • Bispecific formats could simultaneously target C10orf82 and functionally related proteins

• Computational Design Opportunities:

  • Machine learning approaches can predict optimal antibody sequences for specific applications

  • Biophysics-informed models enable the design of antibodies with customized specificity profiles

  • These models can generate novel antibody variants beyond those observed in experimental libraries

• High-Throughput Selection Methods:

  • Phage display with synthetic libraries offers increased control over antibody properties

  • Deep sequencing analysis of selection experiments helps identify optimal binding variants

  • Systematic selection against multiple related epitopes can generate antibodies with precise specificity

These engineering approaches hold significant potential for developing next-generation C10orf82 antibodies with enhanced specificity, affinity, and functional properties to support increasingly sophisticated research applications.