Choosing the Right Spatial Proteomics Tool: Comparing Immunofluorescence, Imaging Mass Cytometry, and MIBI – A Handy Guide
The field of immuno-oncology relies on understanding the interplay between tumor cells and the immune system within the tumor microenvironment (TME). Traditional methods like immunohistochemistry (IHC) and immunofluorescence (IF) have limitations. Here, we explore advanced spatial proteomics tools and their practical benefits for lab workflows and sample preservation.
Current Limitations of Traditional Methods
Immunohistochemistry (IHC):
- Single Protein Analysis: Requires multiple tissue sections for a comprehensive protein profile.
- Increased Workload: Multiple sections increase processing time.
- Sample Depletion: Uses up valuable tissue samples.
- Morphological Artifacts: Repeated sectioning can distort tissue integrity.
Immunofluorescence (IF):
- Limited Multiplexing: Visualizes 6-10 markers per section.
- Spectral Overlap: Challenges in distinguishing signals from multiple fluorophores.
- Autofluorescence: Background signals complicate interpretation.
To overcome these limitations, researchers are increasingly exploring metal isotope tagging for TME proteomic analysis.
Metal Isotope Tagging for Enhanced Multiplexing
Antibodies tagged with metal isotopes are used with mass spectrometry for protein identification and quantification. The distinct isotopic masses provide clear, unambiguous signals for each targeted protein in specific tissue locations.
Emerging Technologies:
- Imaging Mass Cytometry (IMC) (Laser ablation+ToF mass spectrometry)
- Multiplex Ion Beam Imaging (MIBI) (Focused ion beam + ToF mass spectrometry)
Both of these emerging technologies capitalize on the metal tagging approach. By eliminating the complexities associated with fluorescence, they offer significant advantages:
- Enhanced multiplexing: These techniques enable the visualization of dozens of proteins within a single tissue section, providing a more comprehensive picture of the TME proteome.
- Streamlined workflows: The ease of creating custom antibody panels eliminates the time-consuming optimization and cumbersome workflow steps required for IHC and IF.
Distinguishing Features of IMC and MIBI
While targeted mass spectrometry-based imaging platforms offer substantial benefits over IHC and IF, by utilizing a high precision, focused ion beam, MIBI stands out due to some key advantages:
- Spatial resolution: MIBI offers a superior spatial resolution (as low as 250 nm) [1] compared to alternative commercially available (of 1 um), targeted mass spectrometry imaging platforms.
- Sample re-interrogation: The use of a laser in IMC consumes the entire tissue while the focused ion beam approach utilized by MIBI allows for repeated analyses of the same sample.
The table below summarizes key features of various spatial proteomic techniques.
| IHC | IF | IMC | MIBI | |
|---|---|---|---|---|
| Plex | 1-2 | 8 | 40+ | 40+ |
| Simple staining workflow | yes | no | yes | yes |
| Subcellular structure imaging | yes | yes | no | yes |
| Tissue preservation | yes | yes | no | yes |
Why MIBI Stands Out as the Optimal Spatial Proteomic Solution
- Exceptional Multiplexing: Analyze over 40 proteins simultaneously.
- High-Resolution Imaging: Preserves subcellular detail.
- Sample Preservation: Enables re-interrogation of samples for deeper exploration.
MIBI allows researchers to delve deeper into the TME and accelerate therapeutic discovery, saving sample tissues, time and costs.
Unleash the transformative power of MIBI in your research. Learn more about the MIBIscope instrument and the MIBIplus service offering.
References
- Leeat Keren et al., MIBI-TOF: A multiplexed imaging platform relates cellular phenotypes and tissue structure. Sci. Adv. 5, eaax5851 (2019). DOI: 10.1126/sciadv.aax5851