Synthesis, Characterization, and Applications of a Superior Dendrimer-Based Polymer for Multiplexed Ion Beam Imaging Time-of-Flight Analysis

High-dimensional single-cell mass spectrometric imagingtechniquessuch as multiplexed ion beam imaging by time-of-flight mass spectrometry(MIBI-TOF), imaging mass cytometry (IMC), and flow cytometry-basedCyTOF utilize antibodies conjugated to linear metal-chelating polymers.Here, we report on the synthesis and characterization of a dendrimer-basedpolymer and its utilization in tissue imaging using MIBI-TOF. We comparedthe staining performance in FFPE tissue of antibodies for lineage-specificimmune proteins (CD20, CD3, CD45, FoxP3) that were conjugated withdendrimer or linear polymer. Staining of serial tissue sections withdendron-conjugated and linear-polymer-conjugated antibodies revealedcomparable avidities of dendrons and linear polymers with log(2) (ratio of mean positive pixel intensity of staining for linearpolymers to dendrons) within the range & PLUSMN;0.25. Interestingly,dendron-conjugated antibodies were observed to have some advantagesover linear polymer-conjugated antibodies. For example, tissue stainingof a nuclear protein, FoxP3 with dendron-conjugated antibodies showednotably less background staining than that of linear-polymer-conjugatedantibodies. Additionally, dendron-conjugated antibodies did not exhibitoff-target cytosolic binding in neural tissue typically observed whenusing linear polymer conjugates. Taken together, this work providesa versatile framework for using third-generation dendron-conjugatedantibodies with improved staining over conventional linear polymers.

Automated summary

This study introduces a synthesis method of a dendrimer-based polymer and its application in MIBI-TOF tissue imaging. By comparing the staining performance of antibodies conjugated with dendrimers or linear polymers in FFPE tissue, it was found that dendrimers and linear polymers have comparable avidities. Moreover, dendrimers have advantages such as less background staining and no off-target binding in neural tissue. Overall, this research provides a versatile framework for using third-generation dendrimer-conjugated antibodies with improved staining compared to conventional linear polymers.