HEK293 Human CALCRL/RAMP1(CGRP) CRE-Luc Cell Line

For research use only. Not intended for human or animal clinical trials, therapeutic or diagnostic use.

Figure 1. Recombinant CALCRL/RAMP1 CRE-Luc HEK293 constitutively expressing CALCRL.

The median FL4-A signal reaches 8.88E4, while the parental HEK293 control shows a median value of 586. The clear separation of two peaks verifies successful stable expression of CALCRL in this cell clone.

Figure 2. WB of CALCRL/RAMP1 CRE-Luc HEK293

Lane 1: parental HEK293 with no detectable RAMP1 band; Lane 2: CALCRL/RAMP1 CRE-Luc HEK293 clone shows clear RAMP1 band near 17 kDa, confirming RAMP1 expression.

Figure 3. Dose Response of Agonists in CALCRL/RAMP1 CRE Luc Reporter HEK293 Cells (C1)

The EC50 value of α-CGRP is 64.2 pM, and β-CGRP shows an EC50 of 361.2 pM. The robust signal response confirms the cell line is functionally intact and suitable for HTS and pharmacological profiling.

Figure 4. Inhibtion of αCGRP/βCGRP  induced Reporter Activity by CGRP Neutralizing Antibodies in CALCRL/RAMP1 CRE-Luc HEK293(C1).

Figure 5. HTRF cAMP Assay with CALCRL/RAMP1 CRE-Luc HEK293(C1)

α-CGRP yields an EC50 of 0.06 nM, while β-CGRP shows an EC50 of 0.36 nM. The dose-dependent cAMP inhibition response confirms the intact GPCR signaling function of this cell line for pharmacological assays.

Cell Resuscitation
1)Rapidly thaw the frozen cells in a 37 °C water bath for approximately 60 seconds. Once thawed (which may take slightly less or more than 60 seconds), immediately transfer the cell suspension from the cryovial into a 15 mL centrifuge tube containing 10 mL of pre-warmed HEK293 Human CALCRL/RAMP1 CRE-Luc Cell Line complete culture medium.
2)Centrifuge cells at 1000 rpm for 5 min to remove medium, then resuspend cells in 5 mL of pre-warmed complete medium.
3)Transfer the cell suspension into a T25 culture flask and incubate at 37 °C with 5% CO₂.
4)After approximately 24–36 hours, replace the medium or passage the cells to remove non-adherent dead cells.

Subculturing procedure
1)When the cell density reaches the appropriate confluency for passaging, wash the cells with PBS, then add 1 mL trypsin to detach the cells. When more than 80% of the cells detach upon gently tapping the culture flask, add complete culture medium to terminate digestion. Gently pipette to obtain a single-cell suspension, transfer to a 15 mL centrifuge tube, and centrifuge at 1000 rpm for 5 minutes.

2)Discard supernatant after centrifugation. Resuspend cells in fresh medium to a single-cell suspension and transfer to a new culture flask for continued growth.

Cell Freezing
After trypsinization and centrifugation of cells from each T75 flask or 10 cm culture dish, discard the supernatant. Add 2 mL of cryopreservation medium (90% FBS + 10% DMSO), gently resuspend thoroughly, and aliquot into two cryovials. Immediately place the cryovials into a controlled-rate freezing container (e.g., Nalgene 5100-0001), fill with isopropanol to the indicated level, and store at −80 °C. After 24 hours, transfer the cryovials to liquid nitrogen for long-term storage.