Moving Forward with Multiplexing

The EVOS lineup from Thermo Fisher Scientific emphasizes sophisticated simplicity, offering usability for scientists – even those with less experience, while still producing high-resolution microscopy images. The new EVOS S1000 Spatial Imaging System uses spectral technology – allowing users to capture images of tissue samples with multiple markers in a single step. Here, The Pathologist speaks with Chris Langsdorf and Adyary Fallarero at Thermo Fisher Scientific to learn more about the EVOS S1000 Spatial Imaging System – and the future of multiplexed tissue samples.

How does spectral technology overcome some of the barriers to multiplexing multiple biomarkers in a single sample?
 

The workflow for spatial biologists performing multiplexing and spatial imaging is roughly divided into three phases. The first is fluorescence labeling – finding the right combination of targets – antibodies and fluorophores to label the tissue. We’re working on simplifying this process to help users prepare samples more effectively. Secondly, the imaging process itself can be slow, difficult, and sometimes lacks sufficient resolution. The goal is to make it simpler for users and speed up this process while maintaining high resolution. And thirdly, analyzing the data of a single tissue sample remains complicated. Extracting quantitative data from images (what we call “feature extraction”) is an area we’re continuously investigating so that we can improve the process for users.

Traditional fluorescence microscopy methods typically allow imaging of up to four targets. However, for spatial biology, four targets is insufficient to truly understand cellular diversity. To overcome that barrier, researchers now use fluorophore-conjugated primary antibodies or amplification strategies to study a larger number of targets. The shift from traditional methods to these new approaches is significant, and researchers need support to adapt to these changes effectively.

What are the benefits of the multiplexing approach?
 

A multiplexed approach in spatial biology provides a much deeper understanding of heterogeneous cell populations while keeping the spatial context of molecular data intact. For example, when comparing healthy and cancerous tissues, we can detect distinct patterns and cell types that may be likely missed with fewer markers. This is particularly important for improving our understanding and developing more accurate biomarkers for tracking disease progression, developing more effective therapies, and achieving improved clinical trial outcomes.

This approach also makes use of many of the same tools as in standard fluorescence microscopy, such as fluorophore-conjugated antibodies for detecting antigens and identifying different cell types in a tissue sample. We also use standard reagents, such as streptavidin conjugates for biotinylated antibody detection and DAPI to stain cell nuclei.

How does the EVOS S1000 Spatial Imaging System aid in the visualization of multiplex biomarkers in single tissue samples?
 

Using the EVOS S1000 can be truly advantageous because it enables both traditional color imaging and advanced fluorescent microscopy with high resolution. It has an intuitive, streamlined acquisition software that enables easy protocol set-up and fast scanning capabilities – acquiring 8 protein targets and DAPI in one single round, due to its spectral technology.

In addition, the system is designed to handle a wide range of dyes and spectral acquisitions, allowing it to work with over 30 different dyes, including Alexa Fluor, Alexa Fluor Plus, and Aluora Spatial Amplification Reagents, among others. It offers great flexibility and removes the limitations of using only certain specific fluorophores, which gives more options for labeling and imaging.

What impact will the new system have on lab workflows?
 

The EVOS S1000 improves lab workflows by offering fast scanning with the highest level of imaging with a beginner friendly acquisition software, similarly to all other EVOS systems. Pathologists familiar with traditional staining methods can confidently transition to multiplex fluorescence imaging without needing to become experts, thanks to the system’s ease of use.

We’re focused on making the entire process – from sample labeling to image acquisition – simple and user-friendly, addressing common challenges at every step. And we aren’t stopping there.