Optics Express· 2026Q1
Continuous-Wave SOFISM with SPAD Array Detection
- 0citations
- Q1SCImago
- 2026year
Short summary
A new continuous-wave (CW) super-resolution optical fluctuation imaging microscopy (SOFISM) method using a SPAD array detector achieves microsecond-scale fluorescence fluctuation analysis and improved spatial resolution over standard image scanning microscopy (ISM).
AI-generated from the title and abstract; the full text is not read.
Key points
- Developed a continuous-wave (CW) SOFISM method using a SPAD array detector.
- Alexa Fluor 647 exhibits microsecond-scale fluorescence fluctuations suitable for CW SOFISM.
- Optimal binning time for SPAD detection was determined to be 2.5 μs.
- Demonstrated improved spatial resolution compared to ISM on HeLa cell microtubules.
- The approach is generalizable to other fluorophores like CF568 and AF555.
AI-generated from the title and abstract; the full text is not read.
Abstract
We present a continuous-wave (CW) implementation of super-resolution optical fluctuation imaging with image scanning microscopy (SOFISM) using a pixelated single-photon avalanche diode (SPAD) detector camera. In a scanning-based geometry, SOFISM requires fluorescence fluctuations on timescales compatible with the pixel dwell time while maintaining sufficient signal under confocal excitation. Photoswitchable fluorophores in an aqueous switching buffer containing the glucose oxidase-catalase oxygen-scavenging system (GLOXY) and mercaptoethylamine (MEA) were therefore used to promote blinking and reduce photobleaching. Correlation analysis of sparse-emitter measurements showed that Alexa Fluor 647 (AF647) exhibits microsecond-scale fluctuation dynamics suitable for CW SOFISM and guided the choice of imaging conditions for scanned samples. Frame rate and delay accumulation were optimized from detector-pair subset variability using a mean-to-STD criterion. The optimal binning time was 2.5 $μ$s, underscoring the advantage of SPAD-based detection for resolving the relevant fluctuation dynamics. The method was demonstrated on HeLa cell microtubules labeled with AF647, yielding improved spatial resolution in the constructed SOFISM images compared with the corresponding ISM images. Additional super-resolved images from samples labeled with CF568 and Alexa Fluor 555 (AF555) showed the approach is readily generalizable to other fluorophores. These results establish CW SOFISM as a practical fluctuation-based super-resolution method for confocal microscopy.
The authors' abstract, as published at the source. Optics Express, 2026 · DOI ↗
Continue with a free account
Ask the paper: 3 free questions a day about this paper; save it, get its citation, new summaries every day for your field. Takeaways are Premium.
Continue free on the webSign in with Google or Apple; no card needed. You come back to this paper.
On your phone:
BiophysicsBiochemistry, Genetics and Molecular Biology