HEK-Blue™ hDectin-1b Cells

SEAP Reporter 293 cells expressing the human Dectin-1b gene

HEK-Blue™ hDectin-1b cells have been engineered to express a secreted alkaline phosphatase (SEAP) in response to stimulation by Dectin-1 ligands. 

Dectin-1 is a C-type lectin receptor (CLR) that recognizes β-glucans, glucose polymers found in the cell walls of fungi, and thus plays an important role in antifungal innate immunity. Dectin-1 is alternatively spliced into 2 major isoforms: a full-length A isoform and a ‘stalkless’ B isoform, which do not induce the same response to soluble and particulate β-glucans [1].

HEK-Blue™ hDectin-1b cells are derived from the human embryonic kidney HEK293 cells which do not naturally express Dectin-1. We have engineered HEK-Blue™ cells that stably express high levels of human Dectin-1b isoform and genes involved in the Dectin-1/NF-κB/SEAP signaling pathway. Levels of SEAP are readily measurable in real-time by a simple colorimetric assay with HEK-Blue™ Detection or QUANTI-Blue™ . In contrast to cells expressing the human Dectin‑1a gene, HEK‑Blue™ hDectin‑1b cells do not respond to soluble β-glucans, such as laminarin and WGP soluble.

To identify Dectin-1-specific responses, we recommend to use HEK-Blue™ Null1-v cells as a control cell line.

Read our review on Dectin-1 & β-glucans.

Read our review on Dectin-1 & inflammasomes.

Reference:

1. Heinsbroek S.E. et al., 2006. Expression of functionally different Dectin-1 isoforms by murine macrophages. J Immunol. 176: 5513.

Figures

NF-κB responses of HEK-Blue™ hDectin-1a and -1b and HEK-Blue™ Null I-v cells (control cell line) to Dectin-1 ligands. Cells were incubated with particulate ligands such as Zymosan (10 μg/ml), WGP dispersible (100 μg/ml) and HKCA (3 x10⁶ cells/ml), or soluble ligands such as Laminarin (100 μg/ml), WGP soluble (10 μg/ml) or TDB (10 μg/ml). TNF-α (10 ng/ml) was used as a positive control. After 24h, SEAP activity was assessed in the supernatant using QUANTI-Blue™, by reading the optical density (OD) at 630 nm.

Dose-dependent inhibition HEK-Blue™ hDectin-1b responses by Laminarin and WGP soluble. Cells were incubated with particulate ligands such as Zymosan (30 μg/ml) or HKCA (3 x10⁶ cells/ml) and soluble ligands such as (a) Laminarin (starting concentration 100 μg/ml) or (b) WGP soluble (starting concentration 1 mg/ml). After 24h, SEAP activity was assessed in the supernatant using QUANTI-Blue™. Data are presented as the percentage of SEAP activity measured in presence of Zymosan or HKCA without Laminarin or WGP soluble.

Specifications

Required Selective Antibiotics: Puromycin and HEK-Blue™ CLR Selection.

Growth medium: DMEM (4.5 g/l glucose), 10% (v/v) fetal bovine serum (FBS), 100 U/ml penicillin, 100 µg/ml streptomycin, 100 µg/ml Normocin™, 2 mM L-glutamine.

Quality Control:

• HEK-Blue™ hDectin-1b cells have been stimulated by various pathogen recognition receptor (PRR) agonists. As expected, Dectin-1 agonists induced the production of SEAP.
• The cell surface expression of human Dectin-1b (hDectin-1b) in this cell line has been validated using fluorescence-activated cell sorting (FACS).
• The stability of this cell line for 20 passages following thawing has been verified.
• These cells are guaranteed mycoplasma-free.

Contents

• 1 vial containing 3-7 x 10⁶ cells
• 1 ml of puromycin (10 mg/ml)
• 2 x 1 ml of HEK-Blue™ CLR Selection (250X)
• 1 ml of Normocin™ (50 mg/ml)
• 1 pouch of HEK-Blue™ Detection 

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