A Comprehensive Performance Analysis of Fluorescent Dyes
In the quest for clearer, more precise imaging, dye choice plays an important role.
One new study evaluates 18 fluorescent dyes, offering a comprehensive guide for DNA-PAINT applications.
Find out which dyes perform best and explore how Chroma’s optical filters contribute to these insights in a research paper, entitled “The DNA-PAINT palette: a comprehensive performance analysis of fluorescent dyes.”
Here is the Abstract of the paper:
DNA points accumulation for imaging in nanoscale topography (DNA-PAINT) is a super-resolution fluorescence microscopy technique that achieves single-molecule ‘blinking’ by transient DNA hybridization. Despite blinking kinetics being largely independent of fluorescent dye choice, the dye employed substantially affects measurement quality. Thus far, there has been no systematic overview of dye performance for DNA-PAINT. Here we defined four key parameters characterizing performance: brightness, signal-to-background ratio, DNA-PAINT docking site damage and off-target signal. We then analyzed 18 fluorescent dyes in three spectral regions and examined them both in DNA origami nanostructures, establishing a reference standard, and in a cellular environment, targeting the nuclear pore complex protein Nup96. Finally, having identified several well-performing dyes for each excitation wavelength, we conducted simultaneous three-color DNA-PAINT combined with Exchange-PAINT to image six protein targets in neurons at ~16 nm resolution in less than 2 h. We thus provide guidelines for DNA-PAINT dye selection and evaluation and an overview of performances of commonly used dyes.
The following products from Chroma Technology were used in the setup for this study:
- ZET488/10× cleanup filter
- ZET561/10× cleanup filter
- ZET642/20× cleanup filter
- ZT488rdc-UF2 emission filter
- ZT561rdc-UF2 emission filter
- ZT647rdc-UF2 emission filter
- ET525/50 m emission filter
- ET500lp emission filter
- ET600/50 m emission filter
- ET575lp emission filter
- ET705/72 m emission filter
- ET665lp emission filter
- TRF89901-EMv2-NK_Nikon TE2000 Laser TIRF Cube quad-band filter cube
- T570lpxr-UF3 longpass dichroic mirror
- T635lpxr-UF3 longpass dichroic mirror
- ET525/50 m filter
- ET595/44 m filter
- ET706/95 m filter
About Chroma Technology
Chroma Technology is a leading manufacturer and OEM supplier of high performance optical filters used in a wide range of applications, including health care, biomedical research, remote sensing, machine vision, and aerospace. Chroma offers an extensive selection of optical filter sets and filter accessories, as well as custom solutions tailored to customer-specific needs.
For more information or a quotation for this or other Chroma products, contact IL Photonics.