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Programming in situ immunofluorescence intensities through interchangeable reactions of dynamic DNA complexes
Jan Zimak1, Ryan M Schweller, Dzifa Y Duose
1Departments of Bioengineering and Chemistry, Rice University, Houston, Texas 77030, USA.
Insights
Researchers developed DNA complexes to precisely control antibody fluorescence intensity in imaging. This method allows for better visualization of molecular targets and reduces signal interference in multiplexed imaging applications.
Area of Science:
- Biochemistry
- Molecular Biology
- Bioimaging
Background:
- Accurate antibody reporting intensity is crucial for in situ fluorescence imaging.
- Current methods lack precise control, hindering visualization of sparse targets and multiplexed imaging.
Purpose of the Study:
- To develop a controllable method for tuning antibody fluorescence intensity.
- To enable precise control over marker reporting levels for enhanced imaging.
Main Methods:
- Designed linear and branched DNA complexes as interchangeable building blocks.
- Utilized DNA-strand-displacement reactions to assemble fluorescence-reporting complexes.
- Programmed DNA interactions to control dye coupling to antibodies.
Main Results:
- Demonstrated deterministic tuning of dye numbers coupled to antibodies.
- Showcased the ability to increase and balance marker reporting levels.
- Successfully controlled fluorescence intensity within fixed cells.
Conclusions:
- DNA complexes offer a versatile platform for regulating antibody-based fluorescence.
- This approach enhances dynamic detection ranges and minimizes signal crosstalk in multiplexed imaging.
- Provides a novel strategy for precise control in fluorescence imaging analyses.
Abstract:
The regulation of antibody reporting intensities is critical to various in situ fluorescence-imaging analyses. Although such control is often necessary to visualize sparse molecular targets, the ability to tune marker intensities is also essential for highly multiplexed imaging strategies in which marker reporting levels must be tuned both to optimize dynamic detection ranges and to minimize crosstalk between different signals. Existing chemical amplification approaches generally lack such control. Here, we demonstrate that linear and branched DNA complexes can be designed to function as interchangeable building blocks that can be assembled into organized, fluorescence-reporting complexes. We show that the ability to program DNA-strand-displacement reactions between these complexes offers new opportunities to deterministically tune the number of dyes that are coupled to individual antibodies in order both to increase and controllably balance marker reporting levels within fixed cells.
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