The rabbit monoclonal antibody 14B06 specifically binds to the scFv region of a GD2-specific mouse monoclonal antibody (mAb, clone 14G2a). Disialoganglioside GD2 is a glycosphingolipid highly expressed on neuroblastoma, melanoma, and other tumors of neuroectodermal origin, with limited expression in normal tissues. The scFv region of 14G2a has been used to develop GD2-specific chimeric antigen receptor (CAR) T cells utilized in preclinical studies and clinical trials.
Shipping and Storage
⚫ Store undiluted at 2-8°C.
⚫ Avoid prolonged exposure to light.
⚫ Avoid freeze/thaw cycle of the reagent.
⚫ The monoclonal antibody was purified by Protein A.
⚫ The antibody was conjugated with Alexa Fluor 647 under optimum conditions, and unincorporated dye was removed.
(Optional) For Whole Blood Sample
1. Pipette 1 μL Rabbit Anti-Mouse 14G2a Monoclonal Antibody, Alexa Fluor 647 into the bottom of the tube.
2. Add dead cell staining solution and additional fluorochrome conjugated antibodies into the bottom of the tube.
3. Pipette 100 µL of well-mixed, anticoagulated whole blood into the bottom of the tube. Mix gently and thoroughly.
Note Avoid smearing sample down the side of the tube. If the sample remains on the side of the tube, it will not be stained with the reagents.
4. Incubate for 25 minutes in the dark at room temperature (18-25°C).
5. Pipette Red Blood Cell Lysis Solution to the tube. Mix gently and thoroughly. Incubate for 15 minutes in the dark at room temperature (18-25°C).
6. Add 500 μL FACS buffer to the tube. Mix well and centrifuge at 300×g for 5 minutes at room temperature (18-25°C). Aspirate supernatant
completely.
7. Repeat step 6 twice.
8. Add a suitable amount of FACS buffer to resuspend cell and analysis by flow cytometry.
(Optional) For Cell Sample
1. Harvest the cells and wash the cells twice by FACS buffer.
2. Count the cells number and the viability.
3. Resuspend the cell suspension to a concentration up to 1×106 nucleated cells per 100 μL of buffer.
4. Add 1 μL Rabbit Anti-Mouse 14G2a Monoclonal Antibody, Alexa Fluor 647, dead cell staining solution and additional fluorochrome. Mix gently and thoroughly.
5. Incubate for 25 minutes in the dark at room temperature (18-25°C).
6. Add 500 μL FACS buffer to the tube. Mix well and centrifuge at 300×g for 5 minutes at room temperature (18-25°C). Aspirate supernatant
completely.
7. Repeat step 6 twice.
8. Add a suitable amount of FACS buffer to resuspend cell and analysis by flow cytometry.
Tabd 1 Content
