C11orf49 Antibody

What are the validated applications for C11orf49 antibodies?

C11orf49 antibodies have been successfully validated for multiple experimental applications including Western blot (WB), immunohistochemistry (IHC), immunofluorescence (IF), and enzyme-linked immunosorbent assay (ELISA). When selecting an appropriate application, researchers should consider that C11orf49 rabbit polyclonal antibodies have shown reliable detection in various tissue samples with recommended dilution ranges of 1:200-1:2000 for WB, 1:20-1:200 for IHC, and 1:10-1:100 for IF applications . The antibody successfully detects the protein at its observed molecular weight of approximately 31 kDa in Western blots, making it suitable for protein expression studies .

What species cross-reactivity has been confirmed for C11orf49 antibodies?

C11orf49 antibodies have demonstrated confirmed reactivity across human, mouse, and rat samples . This multi-species reactivity makes these antibodies valuable for comparative studies across model organisms. Specifically, positive Western blot detection has been validated in mouse lung tissue and human brain tissue, while positive IHC detection has been confirmed in human colon samples, and IF applications have been validated in HepG2 cells . Researchers should conduct preliminary validation when using these antibodies with other species not explicitly tested by manufacturers.

What is the optimal storage and handling protocol for C11orf49 antibodies?

For maximum stability and activity retention, C11orf49 antibodies should be stored at -20°C without aliquoting . The standard formulation includes PBS with 0.02% sodium azide and 50% glycerol at pH 7.3, which helps maintain antibody integrity during storage . For C11orf49 antibodies conjugated with fluorophores, such as Alexa Fluor 488, storage at 4°C for up to 6 months is recommended to preserve fluorescent signal integrity . Researchers should avoid repeated freeze-thaw cycles and maintain sterile handling conditions to prevent contamination and degradation.

How should positive and negative controls be designed when using C11orf49 antibodies?

Robust experimental design requires appropriate controls when working with C11orf49 antibodies. For positive controls, researchers should consider using mouse lung tissue, human brain tissue, or human colon samples, as these have been validated to express detectable levels of the protein . For cellular studies, HepG2 cells have shown reliable C11orf49 expression suitable for IF applications . For negative controls, researchers should consider:

• Primary antibody omission control

• Isotype control using rabbit IgG at matching concentrations

• Tissue from C11orf49 knockout models (when available)

• Pre-absorption of the antibody with immunizing peptide

These controls help distinguish specific from non-specific binding and validate experimental findings.

What sample preparation protocols optimize C11orf49 detection in different applications?

Sample preparation significantly impacts C11orf49 detection success. For Western blot applications, standard SDS-PAGE followed by transfer to appropriate membrane works effectively with C11orf49 antibodies at 1:500 dilution, as validated with mouse lung tissue . For IHC applications with paraffin-embedded tissues, a 1:50 dilution has been validated for human colon samples . For IF applications, a 1:25 dilution with Rhodamine-labeled goat anti-rabbit IgG secondary antibody has shown good results in HepG2 cells . Researchers should optimize fixation conditions based on their specific tissue or cell type, as overfixation may mask epitopes and reduce antibody binding efficiency.

What is the relationship between C11orf49 and cellular structures or functions?

C11orf49, also designated as CSTPP1, functions as a multi-functional protein with significant roles in regulating nuclear shape and ciliogenesis . Research has revealed that C11orf49/CSTPP1 loss dramatically enhances MAP4 recruitment to microtubules (MTs), which subsequently promotes MT nucleation, polymerization, and nuclear lobulation by facilitating assembly of MTs that penetrate the nucleus . Additionally, C11orf49/CSTPP1 knockout affects actin organization by impairing microtubule association of actin nucleators and MT-actin crosslinkers, leading to cytoplasmic retention of Yes Associated Protein (YAP) and blocking the expression and recruitment of cilium disassembly regulators . This functional profile makes C11orf49 antibodies particularly valuable for studying cytoskeletal regulation and nuclear morphology.

How do C11orf49 protein levels vary across different tissue types?

While comprehensive tissue expression profiling data is limited in the provided search results, experimental validations have detected C11orf49 protein expression in several tissue types. Western blot analysis has successfully detected the protein in mouse lung tissue and human brain tissue . Immunohistochemical staining has confirmed expression in human colon samples . For cellular models, HepG2 cells (human liver cancer cell line) have demonstrated detectable levels suitable for immunofluorescence analysis . This tissue distribution pattern suggests roles in multiple organ systems, though researchers should validate expression levels in their specific tissue of interest before designing comprehensive studies.

What protein-protein interactions has C11orf49 been shown to participate in?

C11orf49/CSTPP1 engages in several key protein interactions that influence its cellular functions. Immunoprecipitation studies have demonstrated that C11orf49/CSTPP1 co-precipitates with four subunits of the tubulin polyglutamylase complex (TPGC) . Specifically, it interacts with TPGS1, LRRC49/CSTPP2, and TTLL1 through its residues 1-225, while robust interactions with TPGS2 or PCM1 require residues 112-377 or 1-111, respectively . Additionally, C11orf49/CSTPP1 shows interaction with α-tubulin through multiple regions, suggesting diverse contact points . These interaction profiles provide valuable targets for co-immunoprecipitation studies using C11orf49 antibodies to investigate cytoskeletal regulation mechanisms.

How can C11orf49 knockout models be validated using C11orf49 antibodies?

Validating C11orf49 knockout models requires careful experimental design using reliable antibodies. When generating C11orf49/CSTPP1 knockout cells via CRISPR/Cas9 gene editing, researchers have successfully used C11orf49 antibodies to confirm protein deletion through Western blot analysis . Additionally, phenotypic validation of knockout models should assess nuclear lobulation and abnormal ciliation, as these are characteristic phenotypes observed in C11orf49/CSTPP1 knockout cells . Reintroduction of C11orf49/CSTPP1 cDNA into knockout cells should rescue these phenotypes, providing functional validation. Importantly, researchers should generate multiple knockout clones (e.g., clone #1 and #5 as referenced) to confirm phenotype specificity and rule out off-target effects .

What methodological approaches can differentiate the functional roles of C11orf49 in nuclear shape control versus cilium disassembly?

Research has demonstrated that C11orf49/CSTPP1's roles in cilium disassembly can be mechanistically distinguished from its function in nuclear shape control . To experimentally differentiate these roles, researchers can:

• Perform domain-specific mutations based on the interaction mapping data showing that C11orf49/CSTPP1 engages with different proteins through distinct domains

• Analyze the effects of truncated protein expressions that preserve specific interaction domains

• Conduct rescue experiments with domain-specific mutants in knockout backgrounds

• Use super-resolution microscopy with C11orf49 antibodies to track subcellular localization during different cell cycle phases

These approaches, combined with appropriate C11orf49 antibodies for detection and validation, allow for functional dissection of this multi-functional protein's diverse cellular roles.

What are common sources of non-specific binding with C11orf49 antibodies and how can they be mitigated?

When working with C11orf49 antibodies, researchers may encounter non-specific binding that complicates data interpretation. Common sources include:

• Insufficient blocking: Optimize blocking with 5% BSA or 5% non-fat dry milk

• Cross-reactivity with similar epitopes: Validate specificity using knockout controls

• Secondary antibody non-specific binding: Include secondary-only controls

• Excessive primary antibody concentration: Titrate antibody dilutions beyond manufacturer recommendations to find optimal signal-to-noise ratio

• Overfixation masking epitopes: Test different fixation protocols and duration

Mitigation strategies include increasing wash duration and frequency, using detergent additives appropriate for the application, and pre-absorbing antibodies with non-specific proteins when possible.

What optimization strategies should be employed when antibody performance varies across experimental replicates?

Variability in C11orf49 antibody performance across experiments may result from several factors. Optimization strategies include:

• Standardizing protein extraction methods for consistent epitope preservation

• Implementing strict temperature control during all incubation steps

• Preparing fresh working dilutions of antibody for each experiment

• Validating lot-to-lot consistency when acquiring new antibody stocks

• Standardizing incubation times and conditions across experiments

• Using automated systems for washing and incubation steps when possible

For quantitative applications, researchers should implement internal loading controls and consider normalizing to total protein levels rather than single housekeeping proteins to account for biological variability.

How might new antibody design technologies improve C11orf49 antibody specificity and versatility?

Emerging antibody design technologies show promise for developing improved C11orf49 antibodies. Recent advances in computational antibody design using RFdiffusion allow for atomically accurate de novo design of antibodies with precise epitope targeting . These approaches can generate antibodies with novel CDR loops that make diverse interactions with target epitopes while differing significantly from sequences in training datasets . For C11orf49 research, these technologies could enable:

• Development of conformation-specific antibodies that recognize distinct functional states

• Engineering of antibodies targeting specific interaction domains with reduced cross-reactivity

• Creation of antibodies with enhanced sensitivity for detecting low expression levels

• Design of bispecific antibodies that simultaneously target C11orf49 and interaction partners

These advanced antibody engineering approaches represent the future direction for more precise and versatile C11orf49 research tools.

What experimental approaches can clarify the role of C11orf49 in disease pathogenesis?

While the search results don't explicitly connect C11orf49 to disease states, its roles in regulating nuclear shape, microtubule dynamics, and ciliation suggest potential implications in diseases involving cytoskeletal dysfunction or ciliopathies. Researchers interested in exploring these connections could:

• Analyze C11orf49 expression in disease tissue microarrays using validated antibodies

• Investigate correlations between C11orf49 mutations/variants and disease phenotypes

• Develop conditional knockout models to assess tissue-specific functions

• Employ proximity labeling techniques with C11orf49 antibodies to identify context-specific interactors

• Use super-resolution microscopy to examine subcellular localization in disease models