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Updated: Feb 8, 2026

Optimized Staining and Proliferation Modeling Methods for Cell Division Monitoring using Cell Tracking Dyes
Published on: December 13, 2012
Multiplexed Division Tracking Dyes for Proliferation-Based Clonal Lineage Tracing
Miles B Horton1,2, Giulio Prevedello3, Julia M Marchingo1,2
1The Walter and Eliza Hall Institute of Medical Research, Parkville 3052, Victoria, Australia.
Abstract:
The generation of cellular heterogeneity is an essential feature of immune responses. Understanding the heritability and asymmetry of phenotypic changes throughout this process requires determination of clonal-level contributions to fate selection. Evaluating intraclonal and interclonal heterogeneity and the influence of distinct fate determinants in large numbers of cell lineages, however, is usually laborious, requiring familial tracing and fate mapping. In this study, we introduce a novel, accessible, high-throughput method for measuring familial fate changes with accompanying statistical tools for testing hypotheses. The method combines multiplexing of division tracking dyes with detection of phenotypic markers to reveal clonal lineage properties. We illustrate the method by studying in vitro-activated mouse CD8+ T cell cultures, reporting division and phenotypic changes at the level of families. This approach has broad utility as it is flexible and adaptable to many cell types and to modifications of in vitro, and potentially in vivo, fate monitoring systems.
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