Recombinant Human TROP-2

Recombinant Human TROP-2 is widely used in research applications because it is a critical biomarker and functional protein involved in tumor growth, metastasis, and therapeutic targeting, especially in cancer biology.

Key scientific reasons to use Recombinant Human TROP-2:

• Cancer Target Validation: TROP-2 is highly expressed in many solid tumors and is directly implicated in promoting tumor cell proliferation, survival, and metastasis through pathways such as calcium signaling, ERK1/2 phosphorylation, and IGF2-IGF1R-Akt activation. Recombinant TROP-2 enables in vitro studies to validate its role as a therapeutic target and to screen for inhibitors or antibodies.

• Antibody and ADC Development: Recombinant TROP-2 is essential for generating and characterizing monoclonal antibodies and antibody-drug conjugates (ADCs) that specifically target TROP-2-positive cancer cells. It allows for precise binding assays, epitope mapping, and affinity measurements, which are critical for therapeutic development.

• Functional Assays: Using recombinant TROP-2 in cell-based assays helps elucidate its role in cell cycle regulation, apoptosis inhibition, and stem cell maintenance. It is also used to study downstream signaling events and resistance mechanisms to targeted therapies.

• Diagnostic Applications: Recombinant TROP-2 serves as a standard or control in immunoassays (e.g., ELISA, Western blot) for quantifying TROP-2 expression in clinical samples, aiding in patient stratification and treatment planning.

• Drug Screening: It provides a platform for high-throughput screening of small molecules, peptides, or biologics that modulate TROP-2 activity, facilitating the discovery of new cancer therapeutics.

• Mechanistic Studies: Recombinant TROP-2 allows researchers to dissect its interactions with other proteins (e.g., IGF IIR, EGF receptor) and to study its biochemical properties, post-translational modifications, and structure-function relationships.

Summary of scientific applications:

• Target validation and mechanistic studies in oncology
• Development and characterization of therapeutic antibodies and ADCs
• Functional assays for cell proliferation, apoptosis, and signaling
• Diagnostic assay standardization
• Drug screening and resistance studies

Using recombinant human TROP-2 ensures experimental reproducibility, specificity, and relevance to human disease models, making it indispensable for translational cancer research and therapeutic development.

You can use recombinant human TROP-2 as a standard for quantification or calibration in your ELISA assays, provided it is of high purity and its concentration is accurately determined. This is a common and accepted practice in quantitative ELISA protocols.

Key considerations and best practices:

• Purity and Characterization: The recombinant TROP-2 protein should be highly purified (typically >95% by PAGE or HPLC) and well-characterized, with a known concentration.
• Standard Curve Preparation: Prepare a standard curve using serial dilutions of the recombinant TROP-2 in the same buffer as your samples to ensure accurate quantification.
• Lot-to-Lot Variation: Be aware that different lots of recombinant protein may have slight differences in immunoreactivity or mass, which can affect quantification. It is best practice to calibrate each new lot against a reference standard if available.
• Carrier Proteins: Some recombinant proteins are supplied with carrier proteins (e.g., BSA) to enhance stability. If your ELISA is sensitive to carrier proteins, use a carrier-free formulation.
• Validation: Validate the standard curve in your specific ELISA system to ensure linearity, sensitivity, and reproducibility. The standard should be run in parallel with your samples in every assay.
• Compatibility: Ensure that the recombinant TROP-2 standard is recognized by the antibodies used in your ELISA (i.e., the epitopes are present and accessible).

Summary Table: Recombinant TROP-2 as ELISA Standard

In summary: Recombinant human TROP-2 is suitable as a standard for ELISA quantification if it is pure, well-characterized, and validated in your assay system. Always prepare a fresh standard curve for each assay and validate the performance with your specific reagents and samples.

Recombinant Human TROP-2 has been validated for several key applications in published research, primarily in the context of cancer biology and therapeutic development.

Validated Applications:

• Antibody-Drug Conjugate (ADC) Development: Recombinant TROP-2 is widely used as a target antigen for the development and validation of ADCs, such as sacituzumab govitecan and other investigational agents. These ADCs have shown efficacy in preclinical and clinical studies against various solid tumors, including triple-negative breast cancer (TNBC), non-small cell lung cancer (NSCLC), and HR+/HER2- breast cancer.

• CAR-T Cell Therapy: Recombinant TROP-2 has been used to validate the specificity and cytotoxicity of TROP-2-targeted CAR-T cells in vitro and in xenograft mouse models. These studies demonstrate robust antitumor activity and help assess potential on-target, off-tumor toxicity.

• Bioassays and Functional Studies: Recombinant TROP-2 protein is used in bioassays to study its role in intracellular calcium signaling, cell adhesion, and tumor growth. It serves as a standard for ELISA, Western blot (WB), and SDS-PAGE to detect and quantify TROP-2 expression.

• Immunohistochemistry (IHC): Recombinant TROP-2 is used to validate antibodies for IHC, enabling the detection of TROP-2 in normal and neoplastic tissues, which is critical for diagnostic and research purposes.

• Cancer Stem Cell and Tumorsphere Assays: Recombinant TROP-2-targeting agents have been shown to reduce cancer stem cell populations and tumorsphere formation in vitro and in vivo, supporting its role in cancer stem cell biology.

• Protein-Protein Interaction Studies: Recombinant TROP-2 is used to investigate its interactions with other proteins, such as IGF IIR, and its involvement in signaling pathways like IGF2-IGF1R-Akt, which are relevant to tumor progression and drug resistance.

Summary Table of Applications

Additional Context:

• Recombinant Human TROP-2 is typically expressed in systems such as wheat germ or mammalian cells, and is available in various tagged formats (e.g., Fc chimera, His-tag) to facilitate purification and detection.
• Its overexpression in multiple epithelial cancers makes it a valuable tool for both basic research and translational studies focused on cancer therapeutics.

If you require protocol details or specific assay recommendations for recombinant TROP-2, please specify the intended application.

To reconstitute and prepare Recombinant Human TROP-2 protein for cell culture experiments, dissolve the lyophilized protein in sterile PBS or distilled water to a recommended concentration (commonly 500 μg/mL), then gently mix and further dilute in cell culture medium containing a stabilizer such as BSA or FBS if required for your application.

Step-by-step protocol:

• Equilibrate materials: Allow the vial and reconstitution buffer (PBS or distilled water) to reach room temperature before opening.
• Centrifuge the vial: Briefly centrifuge or tap the vial to collect all lyophilized powder at the bottom.
• Add buffer: Add the recommended volume of sterile PBS (for carrier-free formulations) or distilled water to achieve the desired concentration (e.g., 500 μg/mL for TROP-2).
• Gentle mixing: Avoid vortexing. Gently swirl or pipette up and down to dissolve the protein. Let stand at room temperature for 15–30 minutes to ensure complete dissolution.
• Dilution for cell culture: After initial reconstitution, dilute the protein in cell culture medium or PBS containing a carrier protein (e.g., 0.1% BSA, 10% FBS, or trehalose) to prevent adsorption and stabilize the protein, unless your experiment requires a serum-free or animal protein-free environment.
• Aliquot and storage: Prepare aliquots sized for single use (≥20 μL) to avoid repeated freeze-thaw cycles. Store aliquots at −20°C to −80°C for long-term stability. After reconstitution, the protein is stable at 2–8°C for up to one week.

Additional considerations:

• For serum-free or animal protein-free applications, use trehalose as a stabilizer instead of BSA or FBS.
• Avoid vigorous shaking or vortexing, as this can denature the protein.
• Always consult the product datasheet or certificate of analysis for specific instructions regarding concentration, buffer, and storage conditions for your batch.

Summary Table:

This protocol ensures optimal solubility, stability, and activity of recombinant TROP-2 protein for cell culture experiments.