RAW-Blue™ Cells

Mouse Macrophage Reporter Cell Line

RAW-Blue™ cells are derived from the murine RAW 264.7 macrophages with chromosomal integration of a secreted embryonic alkaline phosphatase (SEAP) reporter construct inducible by NF-κB and AP-1. RAW-Blue™ cells express many pattern-recognition receptors (PRRs), including toll-like receptors (TLRs), NOD-like receptors (NLRs), RIG-I-like receptors (RLRs) and C-type lectin receptors (CLRs).

Therefore, RAW-Blue™ cells can be used as:

• TLR reporter cells: they express all TLRs, with the exception of TLR5, at different levels
• NOD reporter cells: they express NOD2 and, to a lesser extent, NOD1
• RLR reporter cells: they express RIG-I and MDA-5
• CLR reporter cells: they express Dectin-1

The presence of agonists of these PRRs induces signaling pathways leading to the activation of NF-κB and AP-1, and the subsequent production of SEAP.

Levels of SEAP can be easily monitored using the detection medium QUANTI-Blue™.

RAW-Blue™ Cells were incubated with TLR or NOD agonists: TLR2 (HKLM, 1.108 cells/ml), TLR1/2 (Pam3CSK4, 100 ng/ml), TLR2/6 (FSL-1, 100 ng/ml), TLR3 (poly(I:C), 10 μg/ml), TLR4 (LPS-EK, 1 μg/ml), TLR5 (RecFLA-ST, 1 μg/ml), TLR7 (CL075, 300 ng/ml), TLR9 (ODN1826, 10 μg/ml), NOD1 (Tri-DAP, 10 μg/ml), NOD2 (MDP, 10 μg/ml). After 24h incubation, TLR and NOD stimulation was assessed by measuring the levels of SEAP using QUANTI-Blue™.

RAW-Blue™ cells were stimulated with FSL-1 (10 ng/ml), zymosan (10 μg/ml), depleted zymosan (100 μg/ml), HKSC (10e8 cells/ml) or HKCA (10e8 cells/ml) in the presence or absence of 10 μg/ml of an anti-mTLR2 (clone T2.5) or anti-mDectin-1 monoclonal antibody.
After 24h incubation, NF-kB activation was assessed by measuring the levels of SEAP in the supernatant by using QUANTI-Blue™.

Specifications

Antibiotic resistance: G418, Zeocin™.

Growth medium: DMEM, 4.5 g/l glucose, 2-4 mM L-glutamine, 10% (v/v) fetal bovine serum, 50 U/ml penicillin, 50 μg/ml streptomycin, 100 μg/ml Normocin™ supplemented with Zeocin™ selective antibiotic only.

Guaranteed mycoplasma-free.

Contents

• 1 vial containing 3-7 x 10⁶ cells
• 100 μl Zeocin™ (100 mg/ml)
• 1 ml Normocin™ (50 mg/ml)
• 1 pouch of QUANTI-Blue™ (100 ml)

Shipped on dry ice

Details

Expression of TLR, RLR and NOD mRNAs in RAW-Blue™ cells determined by RT-PCR. RT-PCRs were performed using InvivoGen’s Mouse TLR RT-PCR Primer Set, NOD and RLR RT-Primer Pairs.