Organoid Culture QC Overview
In vitro 3D cell cultures, specifically human organoids, have opened new horizons for translational disease research or modelling, regenerative medicine and predictive precision therapies. Today, differentiated organ cell types can be generated in vitro from a variety of stem cells (SCs) to mimic complex tissues while retaining genomic stability and response to drug treatments.
Thus, organoids have a distinct advantage over traditional 2D cultures and hold unprecedented potential for various, innovative applications. The typical organoid workflow includes multiple steps; SCs are initially isolated from specific organs/tissues and undergo differentiation when cultivated in 3D organoid culture systems, these cultures can then be manipulated (e.g. genetic manipulation, labelling etc.) prior to use in downstream assays (Organoid workflow figure). In order to exploit these models for meaningful basic research, disease modelling or drug screening, specific and reliable culture and analysis methods are required, including tightly controlled, optimized and documented steps throughout the organoid workflow beginning with culture quality control.
Currently, characterization and optimization of organoid cultures are limited in their ability to reproducibly form and monitor these 3D cell models as they form and grow overtime.
- Lack validated protocols for reproducible well-to-well organoid formation and low data throughput
- Involve time-consuming and manual process for acquisition of organoid images
- Require analysis using third party software - with limited quantitative information
- Loss of environmental of control during acquisition of organoid images, which may introduce variability in assay results.
The Incucyte® Organoid Culture QC Application is a turnkey solution consisting of a validated 3D cell culture protocol and software module for real-time visualization and label-free objective quantification to optimize organoid culture conditions.
Incucyte® Organoid Culture QC Application
Generate standardized and reproducible organoid data using live-cell imaging and analysis over days, weeks or months over multiple tissue samples.
Accelerate your organoid discovery with an established protocol and integrated analysis, enabling reproducible culture techniques for downstream assay preparation. The Organoid Culture QC Application allows for direct, label-free analysis of organoid size, count and morphology in Matrigel® domes in a physiologically relevant environment using the Incucyte® Organoid Analysis Software Module and Incucyte® Live-Cell Analysis System.
Incucyte® Organoid Workflow
The Incucyte® Organoid Analysis Software module provides automated label-free analysis solutions for the optimization of organoid culture conditions in 24- or 48- well plates (Incucyte® Organoid Culture QC) and to quantify organoid growth and death in a 96-well plate assay format (Incucyte® Organoid Assay).
Key Advantages of Incucyte® Organoid Assay
- Locate and Analyze Organoids Automatically - Visualize and quantify organoids embedded within Matrigel® domes automatically in a physiologically relevant environment.
- Characterize Organoid Formation Kinetically - Obtain continuous, label-free data to optimize culture conditions using the Incucyte® Organoid Analysis Software Module.
- Identify Maturation Phase of Organoid Cultures - Define an optimal period for passaging organoids or extension of cultures, based on integrated morphology metrics.
- Improve Productivity - Enable scientific discovery using an established protocol and integrated analysis of organoids in real time.
Figure 1. Kinetically monitor and quantify organoid differentiation and growth in Matrigel® domes undisturbed inside your incubator. Mouse intestinal and mouse pancreatic organoids were embedded in Matrigel® domes (50 or 100% Matrigel® respectively) in 24-well plates and imaged over 6 - 8 days in an Incucyte. Brightfield videos (2 d post seeding) and time-course profiles of brightfield area show cell type specific organoid differentiation, morphology and growth respectively.
Characterize Organoid Formation Using Real-Time Kinetic Measurements
Automatically analyse organoid size and count of the entire Matrigel® dome over weeks or months to optimize culture conditions using the Incucyte® Organoid Analysis Software Module.
Figure 2. Distinguish suitable conditions for maximal culture expansion using brightfield size measurements. Mouse hepatic organoids were embedded in Matrigel domes (100%) in 48-well plates at multiple seeding densities. Images (5 d post seeding) and time-courses demonstrate that organoid size and growth rate is directly proportional to cell number.
Identify Maturation Phase of Organoid Cultures
Use unbiased assessment of organoid morphology to provide insight into optimal period for passaging or extending cultures.
Figure 3. Define cell-type specific passaging frequency using integrated morphology metrics. Intestinal organoid passaging frequency is driven by indicators of maturation such as budding and accumulation of debris within the lumen. Kinetically quantify these indicators using Incucyte’s Eccentricity (budding) or Darkness (debris accumulation) metrics.
Leverage an established protocol and integrated software for reproducible formation and analysis of organoids embedded in Matrigel® domes in a 24- or 48-well plates.
Figure 5. Guided interface is easy to use for even first-time users. Automated image acquisition and analysis tools provide a ‘set up and walk away’ experience. Visualize organoid differentiation and growth in 24-well or 48-well plates remotely to monitor experimental progress and analyze in real time.