Semiautomated Production of Cell-Free Biosensors
Cell-free synthetic biology biosensors have potential as effective <i>in vitro</i> diagnostic technologies for the detection of chemical compounds, such as toxins and human health biomarkers. They have several advantages over conventional laboratory-based diagnostic approaches, including the ability to be assembled, freeze-dried, distributed, and then used at the point of need. This makes them an attractive platform for cheap and rapid chemical detection across the globe. Though promising, a major challenge is scaling up biosensor manufacturing to meet the needs of their multiple uses. Currently, cell-free biosensor assembly during lab-scale development is mostly performed manually by the operator, leading to quality control and performance variability issues. Here we explore the use of liquid-handling robotics to manufacture cell-free biosensor reactions. We compare both manual and semiautomated reaction assembly approaches using the Opentrons OT-2 liquid handling platform on two different cell-free gene expression assay systems that constitutively produce colorimetric (LacZ) or fluorescent (GFP) signals. We test the designed protocol by constructing an entire 384-well plate of fluoride-sensing cell-free biosensors and demonstrate that they perform close to expected detection outcomes.
Authors
- Nathan D. McDonald (ORCID: https://orcid.org/0000-0003-3650-9493)
- David C. Garcia (ORCID: https://orcid.org/0000-0002-6029-7964)
- Marilyn F. Slininger Lee (ORCID: https://orcid.org/0000-0002-4555-4803)
- Matthew W. Lux (ORCID: https://orcid.org/0000-0002-2773-742X)
- Katherine A. Rhea (ORCID: https://orcid.org/0000-0002-3741-2549)
- Aníbal Arce (ORCID: https://orcid.org/0000-0001-8134-6385)
- Aleksandr E. Miklos (ORCID: https://orcid.org/0000-0001-6375-2304)
- Casey B. Bernhards
- Daniel A. Phillips (ORCID: https://orcid.org/0000-0003-2759-5246)
- Stephanie D. Cole
- Julius B. Lucks (ORCID: https://orcid.org/0000-0002-0619-6505)
- Brenda Wang
- Walter Thavarajah (ORCID: https://orcid.org/0000-0002-6122-2796)
- Michael C. Jewett (ORCID: https://orcid.org/0000-0003-2948-6211)
- Xavier Bower (ORCID: https://orcid.org/0009-0003-3074-6997)
- Steven M. Blum
- Ashty S. Karim (ORCID: https://orcid.org/0000-0002-5789-7715)
- John R. Biondo
- Tyler Lucci (ORCID: https://orcid.org/0009-0005-7324-4799)
- Jennifer A. Lee
- Dylan M. Brown (ORCID: https://orcid.org/0000-0001-8153-7683)
- John P. Davies (ORCID: https://orcid.org/0000-0001-9873-6830)
- Dale L. Perdue
- Jacob T. Mangini
Institutions
- Northwestern University (US)
- United States Department of the Army (US)
- Defense Threat Reduction Agency (US)
- United States Army Combat Capabilities Development Command (US)
- United States Army (US)
- Compass Systems (United States) (US)
- Precise (India) (IN)
- Synthetic Biologics (United States) (US)
- U.S. Army Combat Capabilities Development Command Chemical Biological Center
- University of Maryland, College Park (US)
- Stanford University (US)
Publication Details
- Journal
- ACS Synthetic Biology
- Published
- 2025-03-12
- DOI
- https://doi.org/10.1021/acssynbio.4c00703
- Citations
- 10
- Primary Topic
- Innovative Microfluidic and Catalytic Techniques Innovation
- Type
- article
- Field-Weighted Citation Impact
- 6.74
Funders
- National Science Foundation
- John Simon Guggenheim Memorial Foundation