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Interferon (IFN) is a multifunctional soluble glycoprotein produced by monocytes and lymphocytes, and serves as a key effector molecule in both innate and adaptive immunity. IFN cytokines are classified into three types—Type I, Type II, and Type III—based on the specific interferon receptors they bind to. Type I interferons comprise a diverse group, including IFN-α, IFN-β, IFN-δ, IFN-κ, IFN-ω, IFN-τ, IFN-ε, and IFN-ξ; Type II interferon is IFN-γ; Type III interferon is IFN-λ, which includes IFN-λ1, IFN-λ2, and IFN-λ3. The primary functions of type I interferons are antiviral and antitumor effects, while type II interferons primarily induce the expression of major histocompatibility complex (MHC) antigens and exert immunomodulatory effects; however, their antiviral activity is weaker than that of type I interferons. Type III interferons, discovered in 2003, are a novel class of interferons with broad-spectrum antiviral and immunomodulatory activities.
 
      Upon ligand binding, both type I and type II receptors are activated, leading to the dimerization and rearrangement of receptor subunits. This triggers the activation of associated JAK kinases through autophosphorylation, which in turn activates STAT proteins. Upon phosphorylation, the activated STAT forms homodimers and subsequently translocates to the cell nucleus, where it initiates the transcription of interferon-stimulated genes.
Type I interferon also induces the formation of the ISGF3 complex, which consists of STAT1, STAT2, and IRF9. The ISGF3 complex binds to the ISRE (interferon-stimulated response element), further inducing the transcription of interferon-stimulated genes containing ISREs within their promoters.
 
      Binding of IFN-γ to the IFN-γ receptor leads to tyrosine phosphorylation of STAT1 at the Tyr701 site. The phosphorylated STAT1 homodimer translocates to the cell nucleus and binds to the GAS (IFN-γ activation site) element, thereby inducing the expression of IFN-γ-regulated genes.

ISRE-Luc THP-1 reporter cells are THP-1 cells in which the Luc luciferase reporter gene is regulated and expressed by ISRE. The mechanism by which IFN binds to its receptor and activates ISRE is shown in the figure below.

Figure 1. Schematic diagram of the ISRE-Luc THP-1 cell model

Figure 2. Dose Response of Recombinant Human IFN in ISRE-Luc THP-1(C1).

Cell Passage Procedures

1.This cell line grows in suspension.
2.Upon receipt, cells should be thawed immediately or stored in liquid nitrogen until use.
3.Before thawing, pre-warm the water bath and culture medium to 37 °C, and prepare a small amount of dry ice.
4.Remove the cryovial from storage and transport it to the cell culture laboratory on dry ice.
5.Rapidly thaw the cells in a 37 °C water bath. Once the cells are completely thawed, spray the cryovial with 70% ethanol for disinfection and transfer it to a biosafety cabinet.
6.Add 10 mL of pre-warmed culture medium into a 15 mL centrifuge tube. Transfer the contents of the cryovial into the tube and centrifuge at 1000 rpm for 5 minutes.
7.Carefully discard the supernatant. Resuspend the cell pellet in 5 mL of pre-warmed culture medium by gentle pipetting. Immediately perform cell counting and adjust the cell density to 3–6 × 10⁵ cells/mL based on the counting results, then transfer the cells into a culture flask.
8.Count the cells every 1–2 days. When the cell density exceeds 1 × 10⁶ cells/mL, passage the cells promptly or add fresh culture medium. Maintain the cell density between 2 × 10⁵ and 1 × 10⁶ cells/mL.

Suspension Cell Cryopreservation Procedure:

1.Collect 8 × 10⁶ cells, centrifuge, and discard the supernatant.
2.Add 1 mL of cell freezing medium (90% FBS + 10% DMSO) and gently pipette to mix thoroughly. Transfer the suspension into a cryovial.
3.Immediately place the cryovial into a controlled-rate freezing container (Nalgene 5100-0001), fill with isopropanol up to the indicated level, and store at −80 °C.
4.After 24 hours, transfer the cryovial to liquid nitrogen for long-term storage.