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Prolactin (PRL) is a classic pituitary hormone, primarily synthesized and secreted by the lactotrophs of the pituitary gland. As a pleiotropic protein, PRL functions as both a circulating hormone and a cytokine in numerous physiological processes, including lactation, reproduction, osmoregulation, immune responses, brain function, metabolism, and angiogenesis. Prolactin promotes the secretion of Thyrotropin-Releasing Hormone (TRH), estrogen, and Vasoactive Intestinal Peptide (VIP), while simultaneously inhibiting the secretion of dopamine.

PRL binds to the Prolactin Receptor (PRLR), thereby initiating the corresponding intracellular signal transduction cascades required to exert its biological functions. The Prolactin Receptor and the Growth Hormone Receptor belong to the Type I cytokine receptor superfamily. Upon PRL binding to the PRLR located on the cell membrane, the PRLR dimerizes and activates the associated JAK2 kinase; this triggers autophosphorylation of the kinase itself as well as tyrosine phosphorylation of the receptor, which in turn recruits and phosphorylates STAT proteins. The phosphorylated STATs subsequently form dimers and translocate into the cell nucleus to regulate target genes (such as Cyclin D1), thereby promoting mammary gland development and milk synthesis.

The PRL Effector Reporter Cell model effectively simulates the in vivo signal transduction pathway of PRL; the underlying principle is illustrated in the figure below.

Figure 1. Schematic Diagram of the HEK293 Human PRL Effector Reporter Cell Model

Figure 2. Recombinant hPRL Effector Reporter Cell stably expressing PRLR.

Figure 3. Dose Response of Recombinant Human Prolactin in PRL Effector Reporter Cell (C7).

Figure 4. Inhibition of Human Prolactin-induced Reporter Activity by hPRLR Blocking Ab in PRL Effector Reporter Cell(C7).

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 PRL Effector Reporter Cell 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.