mtmr10 Antibody

What is MTMR10 and what is its biological significance?

MTMR10 is a member of the myotubularin-related protein family, classified as a probable pseudophosphatase. Its biological significance stems from its structural characteristics, particularly the presence of a glutamic acid (Glu) residue instead of the conserved cysteine (Cys) residue typically found in the dsPTPase catalytic loop. This substitution renders MTMR10 catalytically inactive . This property suggests that MTMR10 likely serves as a regulatory protein rather than an enzyme with direct catalytic activity in cellular pathways. Understanding MTMR10's function is essential for researchers investigating phosphatase-related signaling mechanisms, particularly in contexts where inactive phosphatase homologs play important regulatory roles.

Which techniques can MTMR10 antibodies be reliably used for?

MTMR10 antibodies have been validated for several research techniques, including:

• Western Blotting (WB): Commercial antibodies like ABIN2790822 have been specifically validated for this application

• Immunohistochemistry (IHC): Antibodies such as HPA006081 can be used at dilutions of 1:50-1:200

• Immunofluorescence (IF): The recommended concentration range is 0.25-2 μg/mL for optimal results

For best results in these applications, researchers should optimize antibody concentrations for their specific experimental systems, as tissue type and fixation methods can affect antibody performance.

What is the typical reactivity profile of commercial MTMR10 antibodies?

Commercial MTMR10 antibodies show varying reactivity profiles across species. For instance:

• ABIN2790822 demonstrates reactivity with human (100%), mouse (77%), guinea pig (79%), and horse (77%) MTMR10

• HPA006081 is specifically validated for human MTMR10

These reactivity profiles are typically determined through sequence comparison and experimental validation. The percentages indicate the degree of predicted cross-reactivity based on sequence homology between species. Researchers working with non-human models should carefully verify antibody performance in their specific species of interest before conducting extensive experiments.

What are the recommended storage conditions for MTMR10 antibodies?

MTMR10 antibodies should generally be stored at -20°C for long-term preservation. For short-term use (up to one week), storing at 2-8°C is acceptable . To maintain antibody quality:

• Avoid repeated freeze-thaw cycles, which can degrade antibody performance

• Consider aliquoting the antibody upon receipt to minimize freeze-thaw events

• Follow manufacturer guidelines for specific formulations, as some may contain preservatives like sodium azide that require special handling

Proper storage is crucial for maintaining antibody performance across experiments and ensuring reproducible results in long-term research projects.

How can I validate the specificity of MTMR10 antibodies for my experimental system?

Validating MTMR10 antibody specificity requires a multi-faceted approach:

• Positive and negative controls:

  • Use tissue or cell lines known to express or lack MTMR10

  • Consider MTMR10 knockdown/knockout samples as negative controls

• Peptide competition assays:

  • Pre-incubate the antibody with the immunizing peptide (such as the MTMR10 C-terminal peptide "PRRNSLILKP KPDPAQQTDS QNSDTEQYFR EWFSKPANLH GVILPRVSGT" )

  • Compare results with and without peptide competition

• Multiple antibody validation:

  • Use antibodies targeting different epitopes of MTMR10

  • Consistent results across antibodies increase confidence in specificity

• Mass spectrometry verification:

  • For the most rigorous validation, immunoprecipitate with the MTMR10 antibody and verify the pulled-down protein by mass spectrometry

This comprehensive validation strategy helps distinguish true MTMR10 signals from potential cross-reactivity with related proteins, particularly other members of the myotubularin-related protein family.

What approaches can be used to improve MTMR10 antibody performance in challenging tissue types?

When working with challenging tissues or applications, consider these optimization strategies:

• Antigen retrieval modifications:

  • For formalin-fixed tissues, test both heat-induced and enzymatic antigen retrieval methods

  • Optimize pH conditions (commonly pH 6.0 or pH 9.0 buffers) for your specific tissue

• Signal amplification systems:

  • Implement tyramide signal amplification (TSA) for low abundance targets

  • Consider biotin-streptavidin amplification systems while controlling for endogenous biotin

• Antibody concentration optimization:

  • Perform titration experiments across a wider range than manufacturer recommendations (e.g., 0.1-5 μg/mL for immunofluorescence instead of the standard 0.25-2 μg/mL )

  • Include longer incubation times at lower concentrations to improve signal-to-noise ratio

• Blocking optimization:

  • Test alternative blocking reagents (BSA, normal serum, commercial blockers)

  • Consider dual blocking protocols for tissues with high background

These approaches can significantly improve detection of MTMR10 in tissues where standard protocols yield suboptimal results.

What are the considerations for using MTMR10 antibodies in co-immunoprecipitation studies?

When designing co-immunoprecipitation (co-IP) experiments with MTMR10 antibodies:

• Antibody selection:

  • Choose affinity-purified antibodies like ABIN2790822 that have been validated for specificity

  • Consider whether the epitope (e.g., C-terminal) might interfere with protein-protein interactions

• Lysis buffer optimization:

  • Test both stringent (RIPA) and milder (NP-40 based) lysis buffers

  • Include appropriate protease and phosphatase inhibitors

• Cross-linking considerations:

  • For transient interactions, consider using membrane-permeable crosslinkers

  • Optimize crosslinking time and concentration to preserve interactions without creating artifacts

• Pull-down controls:

  • Include IgG control pull-downs matched to the host species of your MTMR10 antibody

  • Incorporate input samples (pre-IP lysate) at appropriate dilutions

• Protein detection strategy:

  • For detection of co-precipitated proteins, use antibodies raised in different host species than the MTMR10 antibody

  • Consider using protein A/G beads that have been pre-cleared with the lysate

These methodological considerations help ensure that detected interactions are specific to MTMR10 rather than artifacts of the co-IP procedure.

What are common issues when using MTMR10 antibodies in Western blotting and how can they be resolved?

Common issues and solutions for Western blotting with MTMR10 antibodies include:

When optimizing Western blots with MTMR10 antibodies, consider that the predicted molecular weight should be verified against the expected size of human MTMR10, and loading controls should be carefully selected to match the abundance level of MTMR10 in your samples.

How can I optimize immunofluorescence protocols for MTMR10 antibodies in different cell types?

Optimizing immunofluorescence for MTMR10 detection requires cell-type specific considerations:

• Fixation method selection:

  • For cytoplasmic localization: Test 4% paraformaldehyde versus methanol fixation

  • For potential nuclear localization: Consider dual fixation protocols

• Cell-type specific permeabilization:

  • Adherent cells: Standard 0.1-0.3% Triton X-100 is usually sufficient

  • Suspension cells: May require gentler permeabilization (0.05% saponin)

  • Primary cells: May benefit from digitonin for selective membrane permeabilization

• Antibody incubation parameters:

  • Start with the recommended range (0.25-2 μg/mL) and adjust based on results

  • Consider longer incubation times (overnight at 4°C) at lower antibody concentrations

  • Test both one-step and two-step detection systems

• Signal-to-noise optimization:

  • Include appropriate blocking peptides to assess specificity

  • Consider signal amplification for low-abundance expression

  • Test multiple mounting media to reduce autofluorescence

These optimizations should be systematically tested and documented to establish a reliable protocol for your specific cell system.

What considerations should be made when designing immunohistochemistry experiments with MTMR10 antibodies?

When planning immunohistochemistry (IHC) experiments with MTMR10 antibodies:

• Sample preparation considerations:

  • Fixation type and duration significantly impact epitope preservation

  • Section thickness affects antibody penetration (5-7 μm optimal for most applications)

  • Fresh versus archival samples may require different protocols

• Staining protocol development:

  • Begin with manufacturer's recommended dilution range (1:50-1:200)

  • Include positive control tissues with known MTMR10 expression

  • Test multiple antigen retrieval methods (citrate buffer pH 6.0, EDTA buffer pH 9.0)

• Detection system selection:

  • Colorimetric (DAB) versus fluorescent detection depends on experimental goals

  • Polymer-based detection systems often provide superior sensitivity compared to ABC methods

  • Multiplex staining requires careful antibody pairing to avoid cross-reactivity

• Quantification approach:

  • Define scoring criteria before experiment execution

  • Consider digital pathology tools for unbiased quantification

  • Include appropriate controls for normalization

By systematically addressing these considerations, researchers can develop robust IHC protocols for MTMR10 detection across various tissue types.

How can structure-based prediction methods be applied to MTMR10 antibody research?

Structure-based prediction methods, similar to the position-specific structure-scoring matrix (P3SM) approach described for influenza virus antibodies , can be adapted for MTMR10 antibody research:

• Application to epitope mapping:

  • Use Rosetta-based predictions to identify structurally conserved regions in MTMR10

  • Model antibody-antigen interactions to predict binding epitopes

  • Prioritize conservation analysis of predicted binding interfaces

• Cross-reactivity prediction:

  • Apply structural homology modeling to predict potential cross-reactivity with related myotubularin family proteins

  • Use structure-prediction scores to identify antibodies with potential off-target binding

• Optimization of recombinant antibodies:

  • Employ computational design to engineer improved MTMR10-binding antibodies

  • Predict structural effects of framework mutations on binding affinity

  • Model HCDR3 loops to optimize antigen recognition

• Validation methods:

  • Combine in silico predictions with experimental validation using techniques like hydrogen-deuterium exchange mass spectrometry

  • Verify predicted structural features through crystallography

This structure-based approach can complement sequence-based methods, potentially identifying structural homologs that might be missed by sequence analysis alone, similar to how the P3SM method identified antibodies with structural similarity despite sequence divergence .

What are the considerations for using MTMR10 antibodies in studying protein-protein interactions?

When investigating MTMR10 protein-protein interactions:

• Experimental design considerations:

  • Select antibodies that don't interfere with known or predicted interaction sites

  • For the C-terminal specific antibody (ABIN2790822) , consider whether C-terminal interactions might be disrupted

• Methodological approaches:

  • Proximity ligation assays provide higher sensitivity than traditional co-localization

  • FRET/BRET approaches can reveal direct interactions in living cells

  • Co-immunoprecipitation with stringently validated antibodies remains the gold standard

• Controls and validation:

  • Include antibodies against known MTMR10 interacting partners as positive controls

  • Use binding-deficient mutants as negative controls

  • Validate key findings with orthogonal methods (e.g., pull-down with recombinant proteins)

• Advanced techniques:

  • BioID or APEX2 proximity labeling can reveal the wider MTMR10 interactome

  • Crosslinking mass spectrometry can map interaction interfaces at amino acid resolution

  • Live-cell imaging with tagged constructs can be used to confirm antibody-based findings

These approaches provide complementary information about MTMR10's interaction network, helping to elucidate its biological function beyond its catalytically inactive phosphatase domain.

How can I integrate MTMR10 antibody-based detection with other omics approaches?

Integrating MTMR10 antibody detection with multi-omics approaches enables more comprehensive insights:

• Proteomics integration:

  • Use MTMR10 immunoprecipitation followed by mass spectrometry to identify interaction partners

  • Compare antibody-based quantification with mass spectrometry-based quantification for validation

  • Identify post-translational modifications that might affect antibody recognition

• Transcriptomics correlation:

  • Compare MTMR10 protein levels (antibody-based) with mRNA expression to identify post-transcriptional regulation

  • Use transcriptomics data to identify contexts where MTMR10 is differentially expressed for targeted antibody studies

• Functional genomics applications:

  • In CRISPR knockout/knockdown validation, use MTMR10 antibodies to confirm protein depletion

  • For overexpression studies, verify protein levels using calibrated antibody-based methods

• Spatial biology approaches:

  • Combine immunofluorescence with in situ hybridization for simultaneous protein and mRNA detection

  • Use multiplexed antibody panels including MTMR10 for spatial proteomics

  • Correlate spatial data with single-cell transcriptomics for comprehensive cellular heterogeneity analysis

This integrated approach leverages the specificity of antibody-based detection while contextualizing findings within broader molecular datasets.

What emerging technologies could enhance MTMR10 antibody applications in research?

Several emerging technologies have potential to advance MTMR10 antibody applications:

• Advanced imaging approaches:

  • Super-resolution microscopy techniques can reveal MTMR10 subcellular localization beyond diffraction limits

  • Expansion microscopy can physically enlarge samples for improved visualization of MTMR10 complexes

  • Live-cell antibody-based imaging using cell-permeable nanobodies

• Single-cell proteomics integration:

  • Antibody-based CyTOF for single-cell MTMR10 quantification across heterogeneous populations

  • Integration with single-cell RNA-seq for multi-modal analysis

  • Spatial proteomics using multiplexed antibody panels including MTMR10

• Synthetic biology approaches:

  • Split-protein complementation assays using MTMR10 antibody-derived binding domains

  • Optogenetic tools coupled with antibody-based detection for dynamic interaction studies

  • Protein degradation technologies (PROTAC, dTAG) validated with MTMR10 antibodies

• Computational enhancements:

  • Machine learning for improved antibody design targeting specific MTMR10 epitopes

  • AI-assisted image analysis for quantification of antibody-based signals

  • Integrative analysis platforms combining antibody-based data with other molecular datasets

These emerging approaches could significantly expand the utility of MTMR10 antibodies beyond traditional applications, enabling more sophisticated investigations into MTMR10 biology.

How might advances in antibody engineering improve MTMR10-targeted research?

Recent advances in antibody engineering offer opportunities for enhanced MTMR10 research:

• Recombinant antibody technologies:

  • Single-chain variable fragments (scFvs) against MTMR10 could improve tissue penetration

  • Bispecific antibodies could simultaneously target MTMR10 and potential interaction partners

  • Intrabodies designed for specific subcellular compartments could track MTMR10 localization

• Affinity and specificity engineering:

  • Computational design approaches similar to those used for influenza antibodies could optimize MTMR10 binding

  • Directed evolution platforms might generate higher-specificity variants

  • Epitope grafting techniques could transfer MTMR10-specific binding sites to alternative scaffolds

• Functionalized antibodies:

  • Photocrosslinking antibodies could covalently capture transient MTMR10 interactions

  • Enzyme-conjugated antibodies could enable proximity-based labeling of the MTMR10 microenvironment

  • pH-sensitive or conformation-specific antibodies could detect functional states of MTMR10

• Production improvements:

  • Yeast or mammalian display technologies could yield antibodies with improved properties

  • Chemically defined recombinant production could enhance batch-to-batch consistency

  • Alternative scaffolds with improved stability might complement traditional antibodies

These engineering advances could address current limitations in MTMR10 antibody research, particularly regarding specificity, sensitivity, and functional applications.