SOS1 Recombinant Monoclonal Antibody

What is SOS1 and why is it significant in cellular signaling research?

SOS1 functions as a critical signaling protein that activates Ras, a central player in cell growth, proliferation, and differentiation pathways. Its role in signal transduction has broad implications for various physiological processes and diseases, making it an important target for research into cancer and other disorders . SOS1 promotes the exchange of Ras-bound GDP by GTP and regulates phosphorylation of MAP kinase MAPK3 in response to EGF. It also serves as a catalytic component of a trimeric complex that participates in signal transduction from Ras to Rac by promoting Rac-specific guanine nucleotide exchange factor activity .

What types of SOS1 recombinant monoclonal antibodies are available for research?

Several types of SOS1 antibodies are available for research purposes, including:

• Mouse monoclonal antibodies such as clone SOS-01 that react with human and mouse samples and are validated for Western Blot and ICC/IF applications

• Rabbit recombinant monoclonal antibodies that show specificity for human SOS1 and can be used in various applications including ELISA and flow cytometry

• Mouse monoclonal IgG2a Kappa (clone 4C1) antibodies that are specifically reactive against human SOS1 and optimized for ELISA, Proximity Ligation Assay, and Sandwich ELISA

• Rabbit recombinant monoclonal antibodies like clone EPR7480 that are knockout-tested and suitable for IHC-P, WB, and ICC/IF with human samples

How are SOS1 recombinant monoclonal antibodies produced?

SOS1 recombinant monoclonal antibody production involves a meticulously coordinated process using recombinant DNA technology and in vitro cloning. The genes encoding both heavy and light chains of the SOS1 antibody are incorporated into expression vectors, which are then transfected into host cells. This facilitates the recombinant antibody's expression within a cell culture environment. Following expression, the antibody undergoes purification from the supernatant of transfected host cell lines through affinity-chromatography purification methods . This process ensures high specificity and batch-to-batch consistency compared to traditional hybridoma-derived antibodies.

What are the validated applications for SOS1 recombinant monoclonal antibodies?

SOS1 recombinant monoclonal antibodies have been validated for multiple experimental applications, with different clones showing utility in various techniques:

The selection of the appropriate antibody depends on the specific experimental requirements, including the detection method and sample type .

How can SOS1 antibodies be used to study protein-protein interactions?

SOS1 antibodies can be effectively employed in proximity ligation assays (PLA) to study protein-protein interactions. This method allows visualization of protein complexes in situ with high specificity and sensitivity. For example, anti-SOS1 mouse monoclonal antibody (clone 4C1) has been used to analyze interactions between FGFR1 and SOS1 in HeLa cells, as well as between CRKL and SOS1 in Huh7 cells . When paired with antibodies against potential interaction partners (such as anti-FGFR1 rabbit polyclonal antibody), each resulting red dot in the assay represents the detection of a protein-protein interaction complex. This approach provides spatial information about where in the cell these interactions occur, with nuclei counterstained with DAPI for reference .

What dilutions are recommended for different experimental applications of SOS1 antibodies?

Optimal dilutions vary depending on the specific application and antibody clone:

• For Western Blot applications: Anti-SOS1 antibody [EPR7480] is recommended at 1/1000 dilution

• For Flow Cytometry: SOS1 Recombinant Monoclonal Antibody (CSB-RA561936A0HU) is recommended at 1:50-1:200 dilution

• For Proximity Ligation Assay: SOS1 Antibody (4C1) is typically used at 1:50 dilution when paired with antibodies against potential interaction partners

For any application, researchers should conduct preliminary dilution series experiments to determine the optimal concentration for their specific experimental conditions, as antibody performance can vary based on sample type, preparation method, and detection system .

What storage conditions are optimal for maintaining SOS1 antibody activity?

For optimal preservation of antibody activity, SOS1 antibodies should be stored according to manufacturer recommendations, which typically include:

• Storage at -20°C or -80°C immediately upon receipt

• Avoidance of repeated freeze-thaw cycles, which can degrade antibody quality and reduce binding efficiency

• For antibodies formulated with glycerol (such as those in phosphate buffered saline with 50% glycerol), the presence of glycerol allows for storage at -20°C without freezing solid, reducing damage from freeze-thaw cycles

• Aliquoting of antibodies upon receipt to minimize freeze-thaw cycles when only a portion is needed for experiments

Following these storage recommendations ensures maintenance of antibody performance throughout its shelf life .

How can researchers validate the specificity of SOS1 antibodies?

Validation of SOS1 antibody specificity is crucial for reliable experimental results and can be accomplished through multiple approaches:

• Knockout validation: Using cell lines with SOS1 gene knockout as negative controls, as demonstrated with the EPR7480 clone tested on wild-type versus SOS1 knockout A431 cell lysates

• Recombinant protein controls: Testing antibody reactivity against recombinant SOS1 protein, as performed with the 4C1 clone

• Epitope verification: Confirming antibody recognition of the intended epitope, such as the synthesized peptide derived from Human SOS1 used for CSB-RA561936A0HU

• Cross-reactivity assessment: Testing against samples from multiple species to verify species specificity claims

• Application-specific validation: Performing control experiments relevant to the intended application, such as testing secondary antibody-only controls for immunostaining or isotype controls for flow cytometry

What are common technical issues when using SOS1 antibodies in flow cytometry?

When using SOS1 antibodies for flow cytometry applications, researchers may encounter several technical challenges:

• Low signal intensity: This may result from insufficient antibody concentration, poor cell permeabilization (since SOS1 is an intracellular protein), or degradation of the antibody. Optimization strategies include testing higher antibody concentrations (within the recommended 1:50-1:200 range), improving permeabilization protocols, and ensuring proper antibody storage .

• High background signal: Can occur due to non-specific binding, insufficient blocking, or overly high antibody concentration. This can be addressed by increasing blocking agent concentration, optimizing antibody dilution, and including appropriate isotype controls .

• Poor cell fixation: Inadequate fixation can affect epitope accessibility and result in inconsistent staining. For optimal results with SOS1 antibodies in flow cytometry, cells should be properly fixed (e.g., with 4% formaldehyde) and permeabilized (e.g., with 0.2% TritonX-100) before antibody incubation .

• Appropriate controls: For accurate interpretation, include unstained cells, isotype controls, and when possible, positive and negative cell populations. For SOS1 flow cytometry, rabbit IgG can serve as an appropriate control antibody at equivalent concentration .

How can SOS1 antibodies be used to investigate the SOS1-RAS signaling axis in cancer research?

SOS1 antibodies offer valuable tools for investigating the SOS1-RAS signaling axis in cancer research through multiple sophisticated approaches:

• Protein complex characterization: Using proximity ligation assays with SOS1 antibodies enables visualization and quantification of SOS1's interactions with upstream receptors (like FGFR1) and downstream effectors in cancer cell lines, providing insight into aberrant signaling mechanisms .

• Intracellular localization studies: Immunofluorescence with SOS1 antibodies allows researchers to track changes in SOS1 subcellular localization in response to stimuli or drug treatments, which is critical for understanding signal transduction dynamics in cancer cells .

• Expression level analysis: Western blotting with validated SOS1 antibodies enables quantitative assessment of SOS1 expression across different cancer cell lines or patient samples, potentially identifying correlations with disease progression or treatment response .

• Therapeutic target validation: In combination with inhibitors targeting the SOS1-RAS interface, SOS1 antibodies can help validate the mechanism of action through competition assays or by monitoring changes in downstream signaling components .

• Protein-protein interaction modulation studies: As highlighted in recent research, nanobodies that modulate SOS1-RAS interactions have been developed for studying this critical complex. SOS1 antibodies can be used alongside these tools to understand how disrupting or stabilizing these interactions affects cancer cell behavior .

What methodologies combine SOS1 antibodies with other tools to study allosteric regulation?

Recent advances have demonstrated sophisticated methodologies that combine SOS1 antibodies with other molecular tools to study allosteric regulation:

• ChILL and DisCO Approach: A novel methodology combining "Cross-link PPIs and immunize llamas" (ChILL) with "Display and co-selection" (DisCO) has been developed to discover diverse nanobodies that either stabilize or disrupt SOS1-RAS protein-protein interactions. SOS1 antibodies can be used in complementary assays to validate findings from these approaches .

• Competitive vs. Allosteric Binding Analysis: SOS1 antibodies can be employed in competition assays to distinguish between directly competitive and allosteric modulators of SOS1 function. This helps identify whether novel compounds bind at the SOS1-RAS interface or at distant sites that allosterically affect this interaction .

• Sandwich ELISA-Based Detection Systems: Advanced sandwich ELISA techniques using SOS1 antibodies as capture antibodies can detect recombinant GST-tagged SOS1 with high sensitivity (detection limit of 0.1 ng/ml), enabling quantitative analysis of SOS1 variants or modified forms in complex samples .

• Structure-Function Analysis: Combining SOS1 antibodies that recognize specific domains with site-directed mutagenesis allows researchers to correlate structural features with functional outcomes, particularly in understanding how SOS1 transitions between inactive and active conformations .

How can researchers use SOS1 antibodies to investigate its role in disorders beyond cancer?

While SOS1 is prominently studied in cancer, its antibodies can be valuable for investigating its role in other disorders:

• Developmental Disorders: SOS1 mutations are associated with Noonan syndrome, a developmental disorder affecting multiple body systems. SOS1 antibodies can be used to study how these mutations affect protein expression, localization, and interaction with other signaling components in patient-derived cells or model systems .

• Cardiovascular Diseases: The RAS pathway, including SOS1, plays important roles in cardiac development and function. Immunohistochemistry with SOS1 antibodies on cardiac tissue can reveal expression patterns relevant to pathological conditions .

• Immune System Regulation: SOS1 functions in T-cell receptor signaling. Flow cytometry with SOS1 antibodies can be used to analyze expression levels or activation-dependent changes in immune cells from patients with autoimmune disorders .

• Neurological Conditions: As a component of growth factor signaling pathways important for neuronal development and function, SOS1 may be relevant to certain neurological disorders. Proximity ligation assays using SOS1 antibodies can reveal altered interaction patterns in neural cells .

• Metabolic Disorders: The RAS-MAPK pathway interfaces with metabolic signaling networks. Western blotting with SOS1 antibodies can assess changes in expression or post-translational modifications in metabolic disease models .

In each of these research areas, it is critical to select SOS1 antibodies validated for the specific application and sample type, and to include appropriate controls for accurate interpretation of results .

How are SOS1 recombinant monoclonal antibodies being used in combination with nanobody technology?

Recent research has demonstrated innovative approaches combining traditional SOS1 recombinant monoclonal antibodies with emerging nanobody technology:

• Complementary targeting strategies: While conventional SOS1 antibodies typically recognize surface-exposed epitopes, nanobodies can access cavities and hinge regions of SOS1. Using both tools provides comprehensive coverage of different structural features and functional states of the protein .

• Validation and characterization workflows: SOS1 antibodies serve as reference standards in validating newly discovered nanobodies that target SOS1-RAS interactions. The conventional antibodies help confirm the specificity of nanobodies identified through ChILL and DisCO approaches .

• Mechanistic investigations: Nanobodies discovered through specialized immunization strategies can be competitive, connective, or fully allosteric with respect to SOS1-RAS interactions. Traditional SOS1 antibodies can help characterize these different modes of action through competitive binding assays and functional studies .

• Therapeutic development pipeline: While nanobodies that modulate SOS1-RAS interactions show promise as therapeutic leads, conventional SOS1 antibodies remain essential research tools for studying the effects of these interventions on downstream signaling pathways .