SpectraPlex by Leica Microsystems is a cutting-edge software integrated with the STELLARIS confocal platform that streamlines and enhances the workflow of 3D high-multiplex imaging. As highlighted in a Nature Portfolio Application Note, SpectraPlex enhances imaging accuracy with real-time unmixing algorithms that minimize artifacts, optimize signal detection, and eliminate iterative imaging cycles. This enables researchers to visualize multiple biological markers in a single session while maintaining sample integrity. Additionally, its innovative Virtual Fridge system streamlines high-multiplex experiments by organizing, storing, and managing fluorophore selections for reproducible panel development.
In another 2024 Nature Portfolio Application Note, Kunz et. al. of Leica Microsystems used SpectraPlex to build a 3D high-multiplex panel of a mouse pancreatic tumor section. The researchers used Biotium’s CF® Dyes to prepare primary antibody conjugates alpha-SMA-CF®568, FoxP3-CF®633, E-cadherin-CF®660C, and CD31-CF®800 as part of a one-shot, 15-plex workflow. This method captured a large set of targets while preserving both tissue structure and molecular integrity, unlike traditional sequential staining that can compromise samples and limit real-time adaptability of imaging strategies.
SpectraPlex also resolved overlapping fluorescence signals using a “Group” approach, staining three tissue sections with subsets of five markers each to create an unmixing matrix for the full 15-plex dataset. This minimized sample waste, enhanced signal separation, and enabled high-fidelity imaging of intricate tumor structures. By eliminating iterative imaging rounds, SpectraPlex avoided sample degradation and allowed the researchers to revisit regions of interest (ROIs) without introducing artifacts.
Biotium’s CF® Dyes complement SpectraPlex by delivering bright, photostable fluorescence with exceptional signal-to-noise, particularly in the near-IR range. Though four out of the 15 dyes used in the application note were CF® Dyes, as demonstrated in Figure 1b of this image from Nature Magazine, CF® Dyes are available in spectrally unique alternatives for all 15 colors. The synergy between SpectraPlex and CF® Dyes enhances spectral separation and ensures reliable, high-quality imaging for advancing cancer immunology research and diagnostics.
Learn more about Biotium’s bright and photostable CF® Dyes, now available in over 40 colors from blue to near-infrared.
Searching for the best CF® Dye that fits with your instrument configuration? Check out our handy CF® Dye Selection Tool to find the optimal CF® Dye for a specific laser/filter combination for microscopy, flow cytometry, or western blotting.