The rabbit polyclonal antibody R19P specifically binds to the scFv region of a CD19-specific mouse monoclonal antibody (mAb, clone FMC63). CD19 antigen is a B-cell specific cell surface antigen, which is expressed in all B-cell lineage malignancies and normal B-cells. The scFv region of FMC63 has been used to develop CD19-specific chimeric antigen receptor (CAR) T cells utilized in clinical trials.
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Item Information |
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| Catalog # | 500014 |
| Size/Concentration | 100 tests |
| Price | $500.00 |
| Antibody Registration Number | AB_2857948 |
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Specification |
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| Host Species | Rabbit |
| Antibody Type | Polyclonal |
| Antigen Purification | Protein A |
| Reactivity | Mouse |
| Immunogen | scFv region of a CD19-specific mouse mAb clone FMC63 |
| Storage Buffer | Aqueous buffered solution containing protein stabilizer and ≤0.05% ProClin 300 |
| Application | FCM |
Shipping and Storage
| Shipping | The product is shipped at 2-8°C. |
| Stability & Storage |
To provide optimal stability of reagent:
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FACS Protocol
- Reagents Required
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- Phosphate Buffered Saline (PBS with a pH ~7.4).
- FACS Buffer (PBS containing 2% of BSA).
- Rabbit Anti-Mouse FMC63 scFv Polyclonal Antibody, Biotin (100 Tests)
- Preparation of Reagents
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- Rabbit Anti-Mouse FMC63 scFv Polyclonal Antibody, Biotin (100 Tests)
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- Protein must be diluted before use for FACS.
- Dilute the protein at a dilution of 1:100 in FACS buffer.
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- Staining Protocol
- Harvest the cells from source and wash the cells once using FACS buffer.
- Perform a cell count and a cell viability count. Cell viability must be ≥ 95% with a cell number above 2×105 live cells.
- In a round-bottom test tube, resuspend cells in 100 µL of diluted Rabbit Anti-Mouse FMC63 scFv Polyclonal Antibody, Biotin for 30 min at 4°C.
- Wash the mixture of cells and antibody 3 times by centrifuge using FACS buffer.
- Resuspend the stained cells in 200 µL PBS per cell sample.
- Transfer the stained cell into a desired flow tube for analyzation on desired Flow Cytometer. The recommended acquisition rate is >10,000 events.
Product Notices
- Since applications vary, optimal titration should be determined by each investigator for each application to obtain optimal results.
- Caution: Antibody solutions containing ProClin 300 should be handled with care. Do not take internally and avoid all contact with the skin, mucosa
and eyes.
Intellectual Product Notices
For Research Use Only. Not for use in diagnostic or therapeutic procedures
- This product is provided under an intellectual property license from Life Technologies Corporation. The transfer of this product is conditioned on the buyer using the purchased product solely in research conducted by the buyer and the buyer must not use this product or its components:
for:- diagnostic, therapeutic or prophylactic purposes; or
- manufacturing or quality assurance or quality control.
- and/or to sell or transfer this product or its components for resale, whether or not resold for use in research.
- For information on purchasing a license to this product for purposes other than as described above, contact Life Technologies Corporation, 5781 Van Allen Way, Carlsbad, CA 92008 USA or outlicensing@thermofisher.com.
- The information disclosed herein is not to be construed as a recommendation to use the above product in violation of any patents. Bioswan will not be held responsible for patent infringement or other violations that may occur with the use of our products.
- Purchase does not include or carry any right to resell or transfer this product either as a stand-alone product or as a component of another product.
- Any use of this product other than the permitted use without the express written authorization of BioSwan Company is strictly prohibited.
Not for resales. BioSwan, the BioSwan Logo and all other trademarks are property of BioSwan Laboratories, Co., Ltd..
FACS Staining Protocol Using Rabbit Anti-Mouse FMC63 scFv Polyclonal Antibody, Biotin
Required Reagents
- Phosphate Buffered Saline (PBS, pH ~7.4).
- FACS Buffer: PBS supplemented with 2% Bovine Serum Albumin (BSA).
- Rabbit Anti-Mouse FMC63 scFv Polyclonal Antibody, Biotin-conjugated (for 100 tests).
Reagent Preparation
- Rabbit Anti-Mouse FMC63 scFv Polyclonal Antibody, Biotin-conjugated
- This antibody must be diluted prior to use in Fluorescence-Activated Cell Sorting (FACS). Prepare a 1:100 dilution in FACS buffer.
Staining Protocol
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Cell Preparation
- Harvest cells from the culture or tissue source, performing a single wash with FACS buffer.
- Conduct a cell count and assess viability to ensure a minimum viability of 95% and a count exceeding 2×10^5 viable cells.
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Primary Antibody Incubation
- Resuspend the cells in 100 µL of the diluted Rabbit Anti-Mouse FMC63 scFv Polyclonal Antibody, Biotin.
- Incubate for 30 minutes at 4°C.
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Washing
- After incubation, wash the cell-antibody mixture three times by centrifugation with FACS buffer to remove unbound antibodies.
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Resuspension
- Following the final wash, resuspend the stained cells in 200 µL of PBS for each cell sample.
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Flow Cytometry Analysis
- Transfer the stained cells into appropriate flow cytometry tubes for analysis.
- Utilize a flow cytometer with a recommended acquisition rate of over 10,000 events for optimal results.
Notes
- The dilution ratio for the antibody is a guideline; it may require optimization based on specific experimental conditions or the expression level of the target antigen.
- The washing steps are crucial for removing non-specifically bound antibodies, thereby reducing background staining and improving the specificity of the staining.
- For samples with lower cell viability, additional optimization of the staining protocol may be required to improve the accuracy of the analysis
Citation List
- Wang, H., Tsao, ST., Gu, M. et al. A simple and effective method to purify and activate T cells for successful generation of chimeric antigen receptor T (CAR-T) cells from patients with high monocyte count. J Transl Med 20, 608 (2022). https://doi.org/10.1186/s12967-022-03833-6
