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Updated: Jun 15, 2025
Author Spotlight: Importance of Single Cell Sorting in Isolating Purified Populations of Mesenchymal Stem Cells
Published on: November 10, 2023
Single cell, Label free Characterisation of Human Mesenchymal Stromal cell Stemness and Future Growth Potential by
Jared M Campbell1, Abbas Habibalahi2, Adnan Agha2
1Graduate School of Biomedical Engineering, University of New South Wales, Sydney, NSW, 2052, Australia. j.campbell@unsw.edu.au.
Aim:
To use autofluorescence multispectral imaging (AFMI) to develop a non-invasive assay for the in-depth characterisation of human bone marrow derived mesenchymal stromal cells (hBM-MSCs).
Methods:
hBM-MSCs were imaged by AFMI on gridded dishes, stained for endpoints of interest (STRO-1 positivity, alkaline phosphatase, beta galactosidase, DNA content) then relocated and results correlated. Intensity, texture and morphological features were used to characterise the colour distribution of regions of interest, and canonical discriminant analysis was used to separate groups. Additionally, hBM-MSC lines were cultured to arrest, with AFMI images taken after each passage to investigate whether an assay could be developed for growth potential.
Results:
STRO-1 positivity could be predicted with a receiver operator characteristic area under the curve (AUC) of 0.67. For spontaneous differentiation this was 0.66, for entry to the cell-cycle it was 0.77 and for senescence it was 0.77. Growth potential (population doublings remaining) was estimated with an RMSPE = 2.296. The Mean Absolute Error of the final prediction model indicated that growth potential could be predicted with an error of ± 1.86 doublings remaining.
Conclusions:
This non-invasive methodology enabled the in-depth characterisation of hBM-MSCs from a single assay. This approach is advantageous for clinical applications as well as research and stands out for the characterisation of both present status as well as future behaviour. The use of data from five MSC lines with heterogenous AFMI profiles supports potential generalisability.
