An Iodinated DAPI‐Based Reagent for Mass Cytometry

Abstract

Multiparametric single‐cell analysis has seen dramatic improvements with the introduction of mass cytometry (MC) and imaging mass cytometry (IMC™). These technologies expanded the number of biomarkers that can be identified simultaneously by using heavy‐isotope‐tagged antibody reagents. Small‐molecule probes bearing heavy isotopes are emerging as additional useful functional reporters of cellular features. Realizing this, we explored the iodination of DAPI to produce a heavy‐atom‐substituted derivative of the commonly used fluorescent DNA stain. Although exhibiting a drastically reduced fluorescence emission profile, I‐DAPI retains strong binding affinity for DNA. I‐DAPI was used to detect cellular DNA in MC and IMC™ assays with comparable efficiency to known Ir‐containing DNA intercalators. This work suggests repurposing well‐known colorimetric stains through simple reactions could be an effective strategy to develop new, functional MC and IMC™ reagents.

Reagents for Mass Cytometry

“…105 In an attempt to address this issue, DAPI was iodinated to produce I-DAPI (Figure 10). 106 This molecule was an effective DNA stain in SMC and IMC applications; however, no resolution of cell cycle was observed. The reason for lack of cell cycle resolution remains a point of speculation.…”

“…125 The authors took advantage of CD45 as a ubiquitous cell membrane protein with abundant expression in PBMCs to perform live-cell barcoding. A twenty-barcode library was prepared by labeling cells with anti-CD45 bearing Pd ( 104 Pd, 105 Pd, 106 Pd, 108 Pd, and 110 Pd) and In ( 113 In and 115 In) using a 6-choose-3 strategy. The In-bearing reagents were generated using Maxpar polymers.…”

“…An additional limitation is that Cd QDots with naturally occurring Cd would interfere with the 110 Pd mass channel often used for cellular barcoding, while 112 Cd could possibly affect the 113 In signal due to spillover (M +1). Note that the 106 Pd and 108 Pd signals are generally not affected because 106 Cd and 108 Cd only have ∼1% natural abundance.…”

“…Figure10. Structures of DNA staining agents used for cell enumeration and defining single cell events in mass cytometry: Ir-and Rh-containing metallo-intercalators102 and I-DAPI 106. …”

Self-Assembling Systems Based on Pillar[5]arenes and Surfactants for Encapsulation of Diagnostic Dye DAPI

“…A further increase in the concentration of Pillar [5]arene 2 leads to the destruction of self-associates of the dye and their interaction with the 2, while maintaining the luminescence of the dye. A significant excess of compound 2 leads to complete solubilization of DAPI with quenching of luminescence [73,74]. molar ratio of 1:1, which is given for visual display of the type of luminescent nanoparticles (Figure 9c).…”