Sensitive Colorimetric Detection Method for Quantifying Activation of Various HEKblue SEAP Producing Cell Lines

August 22, 2026

BlueskyReddit
Scripps Research
Department of Immunology and Microbiology
Graduate Student

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Company:

Invivogen

Product Name:

QUANTI-Blue™ - SEAP Detection Reagent

Catalog Number:

rep-qbs

The QUANTI-Blue reagent is a key component required for quantifying the activation of NfKB in various HEKblue cell lines such as the HEKblue NOD1 / NOD2, HEKblue TLR2,6, HEKblue TLR2,1 and the THP1-Dual Cells. The reagent allows for colorimetric detection of Secreted Embryonic Alkaline Phosphatase released downstream of NFkB activation of the various cell lines, which could be triggered by either TLR2, NOD1, NOD2 agonists in particular.

Experimental Design and Results Summary

Application

Quantification of NFkB activation in various cell lines expressing immune receptors

Starting Material

Supernatant of HEKblue or THP1 cells treated with activating ligands and QB buffer and reagent stock solution.

Tips

Recommend to prepare the working solution fresh every time before use and to store the stock solution vials at -20 throughout.

Results Summary

After setting up the various cell lines that contain this SEAP reporter and incubating with ligand overnight, we detect SEAP release from cells by adding 20 ul of supernatant from wells containing the cells to 180 ul of freshly prepared quantiblue solution (Diluting 1:100 and mixing 2 components QB Buffer and QB reagent in water) in 96 well plate. After adding the supernatant, the plate is incubated at 37C for 30 minutes to 4 hours, and colorimetric detection is performed by measuring OD 620 nm to OD 650 nm on a device capable of doing so.

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Summary

The Good

Very sensitive with good dynamic range; easy to prepare and use. Can be used for high-throughput screening

The Bad

Expensive, not stable for long periods of time after preparing working solution

The Bottom Line

Recommend for detecting immune activation of various cell lines harboring the SEAP genes. Only downside is high cost of reagent and poor half-life of working solution.

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