SFT2D3 Antibody

What is SFT2D3 and what cellular functions does it perform?

SFT2D3 (SFT2 domain-containing protein 3), also known as MGC5391 or vesicle transport protein SFT2C, is a 22 kDa membrane protein involved in intracellular vesicle trafficking. Current research indicates that SFT2D3 primarily participates in the fusion of retrograde transport vesicles derived from endocytic compartments with the Golgi complex . This protein represents an important component of the cellular trafficking machinery, particularly in maintaining Golgi integrity and function. Understanding SFT2D3's role provides insight into fundamental cellular processes related to protein sorting and membrane dynamics, which has implications for both normal cellular function and disease states where vesicular transport is compromised.

What are the known post-translational modifications (PTMs) of SFT2D3 and how might they affect antibody recognition?

SFT2D3 undergoes several key post-translational modifications that can affect antibody epitope accessibility and recognition. According to UniProt data, SFT2D3 (Q587I9) exhibits phosphorylation at multiple sites including tyrosine residues Y11 and Y97, and serine residue S56 . Additionally, the protein undergoes ubiquitination at lysine residues K16 and K211 . These modifications may significantly alter protein conformation and epitope availability. When selecting antibodies for research applications, researchers should consider whether their target epitopes overlap with these modification sites, as phosphorylation or ubiquitination could mask epitopes or alter antibody binding affinity. For experimental designs involving immunoprecipitation or proximity labeling techniques, PTM status becomes particularly important as modifications may influence protein-protein interactions being studied.

What are the most common techniques for detecting SFT2D3 protein expression in research?

Detection of SFT2D3 protein expression typically employs several complementary techniques, with Western blotting (WB) and immunohistochemistry (IHC) being the most widely validated applications . Western blotting provides quantitative information about SFT2D3 expression levels in cell or tissue lysates, while maintaining the ability to confirm specificity through molecular weight determination (expected at 22 kDa). Immunohistochemistry offers valuable insights into the spatial distribution of SFT2D3 within tissues, which has been validated in tissues such as human pancreas . Some antibodies also support flow cytometry applications, allowing for quantitative assessment of SFT2D3 expression at the single-cell level . ELISA-based detection provides another quantitative approach for measuring SFT2D3 in solution. For comprehensive characterization, researchers should consider employing multiple detection methods, each offering unique advantages for understanding SFT2D3 biology from different perspectives.

What criteria should be used to select the most appropriate SFT2D3 antibody for specific experimental applications?

Selecting the optimal SFT2D3 antibody requires careful consideration of multiple factors based on experimental needs. First, evaluate the antibody's validated applications—whether Western blot (WB), immunohistochemistry (IHC), flow cytometry (FC), or ELISA—ensuring they align with your planned techniques . Species reactivity is equally critical; current commercial antibodies show validated reactivity to human and mouse SFT2D3, with predictions for chicken . The clonality of the antibody affects specificity and reproducibility—while polyclonal antibodies like DF10009 offer high sensitivity by recognizing multiple epitopes, they may introduce batch-to-batch variability . For epitope specificity, consider whether your research requires targeting specific domains of SFT2D3 or avoiding regions subject to post-translational modifications. Finally, prioritize antibodies with robust validation data, including positive and negative controls, knockout validation, or independent confirmation methods. Cross-referencing validation data from resources like Antibodypedia can provide confidence in antibody performance before experimental commitment .

How can researchers validate the specificity of SFT2D3 antibodies in their experimental systems?

Validating SFT2D3 antibody specificity requires a multi-faceted approach to ensure reliable experimental results. Implementation of RNA interference (RNAi) represents a powerful validation strategy, as demonstrated in studies where antibody binding was dramatically reduced in cells stably transduced with shRNAs targeting the gene of interest . This approach can be adapted for SFT2D3 by transfecting cells with specific SFT2D3-targeting shRNAs and confirming reduced antibody signal. Complementary to knockdown approaches, overexpression models provide another validation avenue—transfection of SFT2D3 expression vectors should produce enhanced signal intensity. For definitive validation, CRISPR/Cas9-mediated knockout cells serve as the gold standard negative control. Western blot validation should confirm a single band at the expected molecular weight (22 kDa) , while immunohistochemistry should be validated using tissues with known SFT2D3 expression patterns, such as pancreatic tissue . Additionally, peptide competition assays can verify epitope specificity by demonstrating signal reduction when the antibody is pre-incubated with its target peptide.

What are the optimal protocols for using SFT2D3 antibodies in Western blotting applications?

Optimizing Western blotting protocols for SFT2D3 detection requires careful attention to several key parameters. Sample preparation should begin with efficient membrane protein extraction, as SFT2D3 is a membrane-associated protein . Cell lysis buffers containing mild detergents (0.5-1% NP-40 or Triton X-100) supplemented with protease and phosphatase inhibitors are recommended to preserve protein integrity, particularly given SFT2D3's known phosphorylation sites (Y11, S56, Y97) . For gel electrophoresis, 12-15% polyacrylamide gels provide optimal resolution around the 22 kDa range where SFT2D3 migrates. Transfer to PVDF membranes (rather than nitrocellulose) is preferred for this lower molecular weight protein. Primary antibody incubation should follow manufacturer recommendations, but typical dilutions range from 1:300-1:1000 as demonstrated with HeLa cell lysates . Validation studies should include positive control lysates from tissues or cell lines with confirmed SFT2D3 expression, such as HeLa cells . When interpreting results, researchers should expect a single band at approximately 22 kDa, though post-translational modifications might cause slight mobility shifts. For quantitative Western blotting, normalization to appropriate housekeeping proteins and technical replicates are essential for reliable quantification of SFT2D3 expression changes.

How should immunohistochemistry protocols be optimized for SFT2D3 detection in different tissue types?

Optimizing immunohistochemistry (IHC) protocols for SFT2D3 detection across diverse tissue types requires systematic protocol refinement. Antigen retrieval represents a critical step—heat-induced epitope retrieval (HIER) using citrate buffer (pH 6.0) has shown success for SFT2D3 detection in human pancreatic tissue samples . For tissues with high lipid content or dense extracellular matrix, extending retrieval times may improve antibody penetration. Blocking solutions containing both serum (5%) and BSA (3-5%) help minimize background staining, particularly important when using polyclonal antibodies like those commonly available for SFT2D3 . Antibody dilution optimization should start with manufacturer recommendations (typically 1:50-1:200 for SFT2D3 antibodies) and be adjusted through systematic dilution series for each tissue type. Incubation conditions warrant careful consideration—overnight incubation at 4°C often yields superior staining compared to shorter incubations at room temperature. Detection systems should be matched to expression levels, with amplification systems (such as tyramide signal amplification) considered for low-abundance expression. Each new tissue type requires validation with positive and negative controls, ideally including tissues with known SFT2D3 expression patterns. Specific attention should be paid to membrane localization patterns consistent with SFT2D3's known subcellular distribution, with quantification using digital image analysis for comparative studies.

What considerations are important when using SFT2D3 antibodies for immunoprecipitation studies?

Successful immunoprecipitation (IP) of SFT2D3 presents unique challenges that require careful methodological considerations. First, as a membrane-integrated protein of 22 kDa , SFT2D3 requires lysis conditions that effectively solubilize membrane proteins while preserving antibody epitopes. Buffers containing digitonin (0.5-1%) or CHAPS (0.5-2%) often prove more effective than more stringent detergents like SDS for maintaining native protein conformation. Pre-clearing lysates with protein A/G beads is particularly important when using polyclonal antibodies to reduce non-specific binding. When selecting antibodies for IP, researchers should prioritize those specifically validated for immunoprecipitation applications, as not all WB/IHC-validated antibodies perform effectively in IP. Cross-linking antibodies to beads using dimethyl pimelimidate (DMP) can prevent antibody co-elution with the target protein, which is particularly valuable when studying the lower molecular weight SFT2D3 to avoid IgG heavy chain interference during subsequent analysis. For studying SFT2D3 interaction partners, gentler elution conditions (such as competitive elution with immunizing peptide) help maintain protein-protein interactions. Because SFT2D3 undergoes post-translational modifications including phosphorylation and ubiquitination at specific residues , researchers investigating modification-dependent interactions should consider using phosphatase or deubiquitinase inhibitors in lysis buffers. Finally, validation of IP efficiency using Western blot analysis is essential before proceeding to mass spectrometry or other downstream applications.

How can SFT2D3 antibodies be used to investigate vesicular trafficking pathways in cellular models?

SFT2D3 antibodies offer powerful tools for dissecting vesicular trafficking pathways due to SFT2D3's involvement in retrograde transport from endocytic compartments to the Golgi complex . For dynamic trafficking studies, researchers can employ immunofluorescence microscopy in combination with SFT2D3 antibodies and markers for different cellular compartments. Co-localization analysis with Golgi markers (GM130, TGN46), early endosomal markers (EEA1, Rab5), or recycling endosomal markers (Rab11) can map SFT2D3's distribution and trafficking routes. Live-cell imaging approaches can be developed using fluorescently tagged secondary antibodies against SFT2D3 primary antibodies in semi-permeabilized cell systems. For higher resolution analysis, super-resolution microscopy techniques (STED, STORM, PALM) combined with SFT2D3 immunolabeling enable nanoscale visualization of membrane trafficking events. Biochemical fractionation approaches complemented by Western blotting with SFT2D3 antibodies allow quantitative assessment of the protein's distribution across cellular compartments under different experimental conditions. Proximity labeling techniques (BioID, APEX) using SFT2D3 as bait can identify transient interaction partners involved in vesicular transport. For functional studies, researchers can combine siRNA-mediated knockdown of SFT2D3 with tracking of labeled cargo proteins, using SFT2D3 antibodies to confirm knockdown efficiency. The antibodies can also be employed in electron microscopy studies with gold-conjugated secondary antibodies to precisely localize SFT2D3 relative to membrane structures at ultrastructural resolution.

What are the emerging applications of SFT2D3 antibodies in developmental biology and disease models?

SFT2D3 antibodies are emerging as valuable tools for investigating developmental processes and disease mechanisms where vesicular trafficking plays critical roles. In developmental biology, these antibodies enable spatiotemporal mapping of SFT2D3 expression across embryonic stages, providing insights into how membrane trafficking machinery is established during organogenesis. Particularly relevant is the study of polarized secretion in epithelial development, where SFT2D3's role in Golgi-associated retrograde transport may influence apical-basolateral protein sorting. In neurological disease models, where vesicular trafficking defects underlie numerous pathologies, SFT2D3 antibodies can help determine if trafficking alterations contribute to protein aggregation or mislocalization. Cancer research applications are expanding, building on findings that certain antibodies can distinguish cancer-initiating cell populations through differential binding patterns , suggesting potential roles for membrane trafficking proteins like SFT2D3 in tumor biology. For metabolic disorders, the association of SFT2D3 with metabolism research areas points to applications in studying how trafficking defects might impact metabolic pathways. Technically, multiplex immunofluorescence approaches combining SFT2D3 antibodies with disease markers enable correlation of trafficking changes with pathological states. In therapeutic development, understanding SFT2D3 localization in disease states using validated antibodies could identify novel intervention points for trafficking-targeted treatments. Each application requires careful validation of antibody performance in the specific model system, with particular attention to species cross-reactivity limitations.

How can researchers integrate SFT2D3 antibody-based techniques with advanced proteomics approaches?

Integration of SFT2D3 antibody-based techniques with advanced proteomics creates powerful research platforms for comprehensive characterization of trafficking networks. Immunoprecipitation using validated SFT2D3 antibodies coupled with mass spectrometry (IP-MS) enables identification of SFT2D3 interaction partners, providing insights into its functional relationships within the retrograde trafficking machinery. For studying dynamic interactions, proximity-dependent labeling approaches like BioID or APEX can be combined with SFT2D3 antibody validation to map the protein's interactome across different cellular states. Phosphoproteomics analysis following SFT2D3 immunoprecipitation can reveal how phosphorylation at sites Y11, S56, and Y97 regulates protein function and interactions. Similarly, ubiquitinomic analysis focusing on K16 and K211 modification sites can elucidate how ubiquitination affects SFT2D3 stability and trafficking. For spatial proteomics, fractionation of cellular compartments followed by Western blotting with SFT2D3 antibodies allows mapping of the protein's distribution, which can be correlated with proteomic profiles of these compartments. Targeted proteomics approaches using selected reaction monitoring (SRM) or parallel reaction monitoring (PRM) can be developed for sensitive quantification of SFT2D3 and its interactors, with antibody-based enrichment improving detection of low-abundance species. Cross-linking mass spectrometry (XL-MS) following SFT2D3 immunoprecipitation can reveal structural details of protein complexes. When implementing these integrated approaches, researchers should validate antibody specificity through orthogonal methods and carefully optimize protocols to preserve protein interactions while minimizing introduction of artifacts.

What are common challenges in SFT2D3 antibody-based experiments and how can they be addressed?

Researchers working with SFT2D3 antibodies encounter several technical challenges that require systematic troubleshooting approaches. Background signal issues in immunohistochemistry and immunofluorescence often stem from the polyclonal nature of available SFT2D3 antibodies . This can be mitigated through extended blocking (2-3 hours) with 5% BSA and 5% serum from the secondary antibody host species, combined with titration of primary antibody concentrations. For Western blotting applications, membrane proteins like SFT2D3 may transfer inefficiently, resulting in weak signals. Optimizing transfer conditions for lower molecular weight proteins (22 kDa) by using PVDF membranes, methanol-containing transfer buffers, and shorter transfer times at higher current can improve detection. Epitope masking due to post-translational modifications at Y11, S56, Y97 (phosphorylation) or K16, K211 (ubiquitination) may reduce antibody binding. Treatment with appropriate phosphatases or deubiquitinases prior to immunodetection can restore epitope accessibility when these modifications interfere with antibody recognition. Extraction efficiency represents another common challenge, as SFT2D3's membrane localization necessitates effective solubilization. Comparing different detergents (CHAPS, digitonin, DDM) at varying concentrations can identify optimal extraction conditions that preserve epitope integrity. For quantitative applications, normalization becomes complex when housekeeping proteins are affected by experimental conditions. Multiple reference proteins or total protein normalization methods should be evaluated to ensure reliable quantification of SFT2D3 expression changes.

How can researchers optimize antibody dilutions and incubation conditions for maximum signal-to-noise ratio in SFT2D3 detection?

Achieving optimal signal-to-noise ratios for SFT2D3 detection requires systematic optimization of multiple experimental parameters. Antibody titration should be performed methodically across a wide dilution range (typically 1:100 to 1:2000 for Western blotting; 1:20 to 1:500 for immunohistochemistry), creating a standard curve that identifies the inflection point where specific signal begins to decrease while background remains constant . Temperature and duration of primary antibody incubation significantly impact specificity—while standard protocols often recommend 1-2 hours at room temperature, overnight incubation at 4°C typically provides superior results for membrane proteins like SFT2D3, allowing for equilibrium binding while minimizing non-specific interactions. Blocking buffer composition warrants careful optimization, with comparison of different blocking agents (BSA, non-fat dry milk, normal serum, commercial blocking solutions) at various concentrations (1-5%) to identify the optimal formulation that minimizes background without impeding specific binding. Addition of low concentrations of detergents (0.05-0.1% Tween-20 or Triton X-100) to washing and antibody dilution buffers can reduce hydrophobic non-specific interactions, particularly important for membrane-associated proteins like SFT2D3. Secondary antibody selection and dilution equally impact signal quality—highly cross-adsorbed secondary antibodies at carefully optimized dilutions (typically 1:1000-1:5000) minimize cross-reactivity. For fluorescent detection methods, extending wash steps (at least 3 washes of 10 minutes each) after secondary antibody incubation substantially improves signal-to-noise ratios by removing unbound antibodies more effectively.

What strategies help ensure reproducibility when working with SFT2D3 antibodies across different experimental batches?

Ensuring reproducibility with SFT2D3 antibodies across experimental batches requires implementation of rigorous standardization procedures. First, antibody aliquoting upon receipt prevents repeated freeze-thaw cycles that can compromise binding activity; creating single-use aliquots stored at -80°C ensures consistent antibody performance over time. For polyclonal antibodies commonly used for SFT2D3 detection , lot-to-lot variation represents a significant concern, necessitating side-by-side validation of new lots against previous ones using identical samples and protocols before adopting them for ongoing research. Creating and maintaining detailed standard operating procedures (SOPs) that precisely document all experimental parameters—including buffer compositions, incubation times/temperatures, and equipment settings—facilitates consistent protocol implementation across experiments. Incorporation of positive and negative controls in every experiment provides critical reference points; HeLa cell lysates serve as effective positive controls for SFT2D3 detection , while SFT2D3-knockdown samples create valuable negative controls. When absolute quantification is required, standard curves using recombinant SFT2D3 protein enable batch-independent measurement calibration. For image-based analyses, consistent acquisition parameters (exposure times, gain settings) and standardized analysis workflows prevent artificial variation introduced during data collection and processing. Researcher training and cross-validation ensure consistent technique application, particularly for manual steps like washing procedures that significantly impact background levels. Finally, implementing replicate types strategically—technical replicates assess protocol reliability, while biological replicates confirm result generalizability—provides comprehensive reproducibility assessment.

How are SFT2D3 antibodies being used to investigate potential roles in disease mechanisms?

SFT2D3 antibodies are enabling novel investigations into disease mechanisms through multiple research approaches. Comparative expression analysis using immunohistochemistry and Western blotting with validated SFT2D3 antibodies allows researchers to identify altered expression patterns across normal and pathological tissues. This approach has been particularly informative in pancreatic tissue, where SFT2D3 antibodies have demonstrated specific staining patterns . Given SFT2D3's function in retrograde vesicular transport , researchers are exploring how trafficking dysregulation might contribute to diseases characterized by protein mislocalization or secretory defects. Colocalization studies combining SFT2D3 antibodies with markers of different trafficking compartments help map potential trafficking perturbations in disease states. In cancer research, antibody-based approaches are examining whether altered trafficking pathways involving SFT2D3 contribute to malignant phenotypes, building on observations that certain cell-surface targeted antibodies can distinguish cancer-initiating cell populations . For metabolic disorders, SFT2D3's categorization within metabolism research areas suggests potential roles in metabolic dysregulation that can be investigated through correlative studies of SFT2D3 localization and metabolic phenotypes. Mechanistically, immunoprecipitation with SFT2D3 antibodies followed by mass spectrometry enables comparison of interactome changes between normal and disease states, potentially identifying altered protein interactions that drive pathology. These approaches are revealing how trafficking machinery components like SFT2D3 may contribute to disease mechanisms beyond their conventional roles in cellular homeostasis.

What novel antibody engineering approaches might improve SFT2D3 antibody performance for challenging applications?

Advanced antibody engineering strategies offer promising avenues for enhancing SFT2D3 antibody performance in demanding research applications. Single-chain variable fragment (scFv) constructs, similar to those developed for spike protein studies , could resolve the preferred orientation challenges sometimes encountered in cryo-EM and other structural biology applications involving membrane proteins like SFT2D3. These smaller antibody fragments provide improved access to sterically hindered epitopes that may occur in the membrane-associated regions of SFT2D3. For super-resolution microscopy applications, where conventional antibody sizes (10-15 nm) limit achievable resolution, nanobodies or aptamer-based detection reagents against SFT2D3 could reduce the distance between fluorophore and target, enhancing spatial precision. Site-specific conjugation technologies enable precise control of fluorophore or crosslinker attachment positions on antibodies, improving functional activity compared to random conjugation methods typically used with polyclonal SFT2D3 antibodies . Recombinant antibody production approaches would address lot-to-lot variability issues inherent to polyclonal antibodies, providing consistent performance across experiments. For multiplexed detection applications, antibody engineering to incorporate orthogonal epitope tags facilitates simultaneous detection of SFT2D3 alongside other proteins of interest. Integrating these advanced antibody technologies into SFT2D3 research workflows could significantly enhance detection sensitivity, specificity, and application versatility beyond what is currently possible with conventional antibody formats.

How might integration of SFT2D3 antibodies with emerging single-cell technologies advance understanding of vesicular trafficking heterogeneity?

Integration of SFT2D3 antibodies with cutting-edge single-cell technologies offers unprecedented opportunities to uncover cellular heterogeneity in vesicular trafficking systems. Single-cell proteomics approaches utilizing SFT2D3 antibodies can reveal cell-to-cell variation in trafficking machinery components that may explain differential responses to trafficking perturbations. Mass cytometry (CyTOF) incorporating metal-conjugated SFT2D3 antibodies enables high-dimensional analysis of trafficking protein expression in correlation with dozens of other cellular markers, providing rich phenotypic context for trafficking heterogeneity. Imaging mass cytometry extends this capability to tissue sections, preserving spatial relationships between cells with different trafficking profiles. Microfluidic platforms for single-cell Western blotting with SFT2D3 antibodies deliver quantitative protein expression data at single-cell resolution, revealing subpopulations with distinct trafficking machinery compositions. For functional characterization, combining SFT2D3 immunostaining with live-cell imaging of vesicular cargo in microfluidic traps allows correlation between trafficking machinery levels and transport kinetics at single-cell resolution. Spatial transcriptomics approaches paired with SFT2D3 immunohistochemistry can connect protein localization patterns with underlying gene expression programs driving trafficking heterogeneity. In developmental and disease contexts, these integrated approaches using validated SFT2D3 antibodies could reveal how trafficking machinery heterogeneity contributes to cell fate decisions or differential disease susceptibility. This technological integration moves beyond population averages to uncover fundamental principles of trafficking system organization and plasticity at unprecedented resolution.

What are the reported sensitivity and specificity metrics for currently available SFT2D3 antibodies?

Performance metrics for commercially available SFT2D3 antibodies demonstrate varying sensitivity and specificity profiles depending on application and target tissue. For Western blotting applications, polyclonal antibodies like DF10009 have demonstrated detection sensitivity sufficient for endogenous SFT2D3 levels in HeLa cell lysates using standard ECL detection systems . Dilution optimization experiments typically recommend working concentrations between 1:300-1:1000 for Western blotting applications, balancing sensitivity with background minimization . For immunohistochemistry applications on formalin-fixed paraffin-embedded (FFPE) tissues, optimal dilutions generally range from 1:50-1:200, with validated staining in human pancreatic tissue sections . Regarding specificity characteristics, several SFT2D3 antibodies have undergone validation through knockout/knockdown approaches that confirm signal reduction following target depletion, similar to methodologies employed for other antibody validations . Cross-reactivity profiles indicate that most commercially available SFT2D3 antibodies demonstrate reactivity against both human and mouse orthologs , with predicted but not extensively validated reactivity against chicken SFT2D3 . Binding affinity data, where available, indicates nanomolar range KD values similar to those reported for other research antibodies (10^-9 to 10^-11 M range) . These metrics provide important benchmarks for researchers selecting SFT2D3 antibodies for specific applications, though comprehensive side-by-side comparisons across the full range of available antibodies remain limited in the published literature.

What epitope mapping data exists for current SFT2D3 antibodies, and how does epitope location affect experimental applications?

Epitope mapping data for SFT2D3 antibodies remains partially characterized in the current literature, with important implications for experimental design. Available SFT2D3 antibodies are typically raised against the full-length protein or specific peptide regions, though precise epitope mapping through techniques like peptide array analysis or hydrogen-deuterium exchange mass spectrometry is not comprehensively reported for most commercial offerings . Based on immunogen information, antibodies raised against different regions of the 221-amino acid SFT2D3 protein (UniProt: Q587I9) likely recognize distinct epitopes . The membrane topology of SFT2D3 creates distinct accessibility considerations—antibodies targeting extracellular/luminal domains are suitable for non-permeabilized immunofluorescence, while those recognizing cytoplasmic domains require permeabilization procedures. Epitopes containing or adjacent to post-translational modification sites (phosphorylation at Y11, S56, Y97; ubiquitination at K16, K211) may show condition-dependent binding, making them valuable for studying modification states but potentially unreliable for total protein quantification. For conformational epitopes spanning membrane-embedded regions, native protein structure preservation becomes critical, limiting application in denaturing techniques like Western blotting while enhancing specificity in native-state applications like immunoprecipitation. When designing experiments involving protein-protein interaction studies, researchers should consider whether antibody epitopes overlap with known interaction interfaces, potentially causing steric hindrance that could yield false-negative results. Multi-epitope approaches using antibody combinations targeting different SFT2D3 regions can provide more comprehensive protein detection and functional insights than single-antibody strategies.

What is known about the cross-reactivity profiles of SFT2D3 antibodies across different species and related proteins?

Cross-reactivity profiles of SFT2D3 antibodies demonstrate both species-specific recognition patterns and potential interaction with structurally related proteins that warrant careful experimental consideration. Current commercial antibodies show validated reactivity against human and mouse SFT2D3 orthologs , with predictive modeling suggesting potential cross-reactivity with chicken SFT2D3 . The basis for this cross-species reactivity lies in sequence conservation analysis—human SFT2D3 shares approximately 82% amino acid identity with its mouse ortholog and 72% with the chicken ortholog, with higher conservation in certain functional domains. Computational prediction models utilizing alignment of immunogen sequences have generated cross-reactivity scores that indicate high confidence (score>80) for chicken reactivity , though experimental validation through Western blotting or immunohistochemistry with chicken samples remains limited in published literature. Within the SFT2 domain-containing protein family, potential cross-reactivity with paralogous proteins SFT2D1 and SFT2D2 represents an important consideration, as these proteins share structural similarities in their SFT2 domains. Comprehensive cross-adsorption studies or immunoblotting against all family members would provide definitive data on paralog specificity, though such systematic analyses are not widely reported for current commercial offerings. For researchers working with less common model organisms, prediction algorithms based on epitope conservation can provide preliminary guidance, but direct validation in the species of interest remains essential. When designing experiments involving novel species or closely related protein families, preliminary validation through recombinant protein controls or knockdown/knockout approaches should be implemented to establish antibody specificity in the specific experimental context.