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Biologic research and disease modeling recapitulate complexity of real tissues

What are organoids?

Organoids are three-dimensional (3D) multi-cellular, microtissues derived from stem cells that are designed to closely mimic the complex structure and functionality of human organs like the lung, liver or brain. Organoids typically consist of a co-culture of cells which demonstrate a high order of self-assembly to allow for an even better representation of complex in vivo cell responses and interactions, as compared to traditional 2D cell cultures.

There are three distinct definitions that differentiate an organoid:

  • It is a 3D biological micro-tissue that contains several types of cells
  • It represents the complexity, organization, and structure of a tissue
  • It resembles at least some aspect of a tissue’s functionality

The importance and advantages of organoids

Organoids are becoming increasingly important in the fields of cancer research, neurobiology, stem cell research, and drug discovery, since they allow for the enhanced modeling of human tissues. Derived from stem cells, organoids can be expanded to a wide range of tissue types including liver, lung, brain, kidney, stomach, and intestine. Because these 3D microtissues mimic in vivo organs, they can provide researchers greater insight into the mechanisms of human development and disease. For example, researchers can grow organoids from mutated cells to understand how specific gene mutations are linked to certain genetic disorders. Organoids can also facilitate the study of infectious diseases and host-pathogen interactions. Finally, the ability to use patient-derived organoids for drug screens and toxicity evaluations is enabling researchers to make further advancements in personalized medicine.

General workflow for organoid research

Due to the growing complexity of organoids and other 3D cellular systems, more sophisticated 3D imaging and analysis techniques are needed to accurately and efficiently characterize these biological assays. Today, automated confocal imaging systems and 3D image analysis software are commonly used to help researchers streamline their workflow and obtain optimal results.

Organoids workflow illustration

Step 1) 2D preculture

Organoids are derived from either primary cells (i.e., intestine, lung, or kidney) or induced pluripotent stem cells. Stem cells are able to differentiate and self-assemble into a variety of tissue-specific organoids.

Step 2) Developing 3D organoids

Typically cells are premixed with Matrigel and droplets are placed into a 24-well plate at room temperature. The plates are then placed into an incubator to form a solid droplet dome. Media is then added for seven or more days to promote cell growth and differentiation into specific tissue (brain, gut, lungs, etc.). Media includes extracellular matrix (ECM), proteins, and different growth factors, which will vary depending on the type of tissue that is being developed.

Step 3) Organoid culture

Organoid culture is a long process and may include several steps with different media. During this process, cell phenotype needs to be monitored (PCR, imaging), typically used for understanding developmental biology and tissues.

Step 4) Monitoring and readouts

Before experiments are conducted, organoids need to be monitored and characterized to ensure that they have the appropriate tissue structure and differentiation. High-content imaging allows for monitoring and visualizing the growth and differentiation of organoids, 3D reconstitution of the structures, complex analysis of organoids structure, cell morphology and viability, as well as the expression of different cell markers.

Step 5) Confocal imaging and 3D analysis

Confocal imaging and 3D analysis of organoids allow visualization and quantitation of the number, size distribution, cell number, cell content, cell viability, volume, as well as quantitation of cell proliferation and expression of specific markers. Characterization of multiple quantitative descriptors of organoids could be used for studying disease phenotypes and compound effects.

Confocal imaging and 3D image analysis of organoids

Organoid structures provide a very useful tool for disease modeling and assessment of compound effects. Automated imaging and analysis of organoids are important for the improvement of quantitative assessment of phenotypic changes in organoids, as well as for the increase of throughput for experiments and tests.


Confocal imaging and 3D image analysis of organoids


Confocal imaging, like the ImageXpress® Confocal system with high-performance lasers and water immersion objectives are especially useful for capturing complexity of 3D biological assays. Unlike typical spheroids, that have appearance of solid objects and limited penetration of light, 3D organoids have a hollow appearance, with a lumen, or cavities inside, and are more easily penetrated by light, allowing “imaging through” multiple microtissues embedded into Matrigel.

Airway Organoid CME analysis

Imaging and Analysis of a 3D Organoid Model. In this short video, Andy Bashford (Imaging Applications Scientist) shows a great example of a 3D Airway Organoid Model along with some interesting ways to get the most out of this type of assay.

High-content analysis tools like the MetaXpress or IN Carta Image Analysis Software, tools allow for the finding and characterization of multiple objects/organoids, either in 2D format (for single plane, or maximum projection images) or in 3D when objects from the multiple planes are connected and reconstituted in 3D space by software. Organoids can be characterized for diameter, volume, shape, intensity of specific marker, or distance to other objects.

Furthermore, individual cells, nuclei, or organelles can be defined and measured within each organoid. That allows count of live and dead cells, or cells with a specific marker also defining volumes and distances between the objects. Numeric values can be counted for each organoid or averaged per well.

Lung organoid cell image gallery

More great resources

Resources for Organoids

Videos & Webinars

Disease modeling in the 21st century: Automated organoid assays with 3D imaging

High-throughput, organoid-derived organ-on-a-chip systems for drug discovery and disease modelling

Developing high-throughput organ-on-a-chip tissue models for drug discovery using high-content imaging

Physiologically-relevant tissue models using a high-throughput Organ-on-a-Chip platform

Physiologically-Relevant Tissue Models Using a High-Throughput Organ-on-a-Chip Platform

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