A quantitative high-throughput chemotaxis assay using bioluminescent reporter cells

Area of Science:

• Biotechnology
• Cell Biology
• Immunology

Background:

• Chemotaxis, the directed cell movement along chemokine gradients, is crucial for immune responses and disease.
• Existing methods for quantifying chemotaxis can be labor-intensive and lack sensitivity.
• A need exists for robust, high-throughput assays to study chemotaxis.

Purpose of the Study:

• To develop and validate a novel biophotonic assay for sensitive and accurate quantitation of cellular chemotaxis.
• To demonstrate the assay's utility in quantifying the chemotactic response of primary human T-cells to specific chemokines.
• To establish a platform for high-throughput screening of chemotaxis modulators.

Main Methods:

• Utilized firefly luciferase (ffLuc) reporter system to generate a biophotonic signal proportional to cell number.
• Quantified cellular migration in a 96-well microplate format using a dedicated chemotaxis instrument.
• Validated the assay by measuring the chemotactic response of ffLuc-expressing primary human T-cells to recombinant chemokines (MCP-1, RANTES, IP-10).

Main Results:

• The biophotonic assay demonstrated superior accuracy, reproducibility, and sensitivity compared to direct cell enumeration.
• Successfully quantified the chemotactic responses of primary human T-cells to MCP-1, RANTES, and IP-10.
• The assay is suitable for analyzing various cell types and biological fluids.

Conclusions:

• The novel biophotonic assay provides a sensitive, accurate, and reproducible method for quantifying chemotaxis.
• This high-throughput compatible system facilitates the screening of large compound libraries for chemotaxis modulators.
• The assay has broad applications in immunology, drug discovery, and cell migration research.