FateView™ flags deviating cell line much earlier than endpoint QC

Cover graphic showing FateView™ identifying PSC line divergence around Day 2–3 during ectoderm differentiation, compared with traditional endpoint analysis at Day 6.

Label-free monitoring detected cell line divergence during ectoderm differentiation by Day 2–3 in a 6-day workflow.  

Differences between pluripotent stem cell (PSC) lines can affect differentiation potential and lead to inconsistent experimental outcomes. In many workflows, these differences are only identified during endpoint analysis, after several days or weeks of a protocol where time and money are invested.  

During a 6-day PSC-to-ectoderm differentiation workflow, FateView™ identified a PSC line deviating from the expected trajectory around Day 2–3. Traditional endpoint immunostaining analysis would not have revealed the issue until Day 6.  

Experimental overview of FateView™ monitoring during PSC-to-ectoderm differentiation

Monitoring ectoderm differentiation using FateView™  

We used a FateView™ assay to compare four PSC lines during an ectoderm differentiation protocol. This assay had previously been developed using immunostaining data from separate experiments to predict the percentage of cells expressing ectoderm marker SOX1 from label-free images.  

Throughout the workflow, live label-free images were uploaded to FateView™ every 4 hours:

Label-free imaging and FateView™ prediction every 4h

To validate FateView™ predictions, cells were fixed every 24 hours for SOX1 staining:

Figure x SOX1 immunostaining was performed every 24 hours during ectoderm differentiation. 

Earlier visibility into differentiation trajectory 

As shown in the figure below, FateView™ SOX1-positive predictions (Solid lines) closely followed the SOX1 immunostaining measurements (X symbols) across the differentiation workflow.  

Line graph comparing FateView™ predictions and SOX1 immunostaining measurements across four PSC lines during ectoderm differentiation. One PSC line diverges from the others around Day 2, enabling earlier detection before endpoint analysis at Day 6.

Cell lines 2-4 showed increasing percentages of SOX1-positive cells over time, consistent with successful ectoderm differentiation.  

Cell line 1 began to diverge around Day 2 and showed reduced ectoderm differentiation relative to the other cell lines. The divergence becomes visible in the FateView™ prediction trajectory around Day 2, whereas traditional endpoint analysis would not have identified this until Day 6. 

Broader applications 

This study demonstrates how FateView™ can identify morphological divergence earlier in a workflow, before endpoint analysis is reached.

Such earlier visibility has the potential to reduce time invested in unsuccessful workflows, support earlier assessment, and improve process consistency.

Beyond ectoderm differentiation, FateView™ can be adapted to identify early differences across cell lines, culture conditions, protocols, or experimental workflows: supporting more confident process development across a wide range of cell systems. 

Learn more about FateView™ and custom assays for specific workflows and experimental readouts. 

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