3D Cell Models

Investigate complex 3D biology using advanced methods
in cellular imaging
 and analysis technologies.

Cellular Imaging and Analysis for 3D Cell Culture Applications and Assays

Development of more complex, biologically relevant, and predictive cell-based assays for compound screening is a primary challenge in drug discovery. The integration of three-dimensional (3D) assay models is becoming more widespread to drive translational biology. Higher complexity cell models have gained popularity because they better mimic in vivo environments and responses to drug treatment. Specifically, 3D cell cultures offer the advantage of closely recapitulating aspects of human tissues including the architecture, cell organization, cell-cell and cell-matrix interactions, and more physiologically-relevant diffusion characteristics. Utilization of 3D cellular assays adds value to research and screening campaigns, spanning the translational gap between 2D cell cultures and whole-animal models. By reproducing important parameters of the in vivo environment, 3D models can provide unique insight into the behavior of stem cells and developing tissues in vitro.

3D cell imaging and analysis workflow

While the development of quantitative assays using 3D cell cultures has become an attractive investigative tool to understand complex biology, challenging 3D cell image acquisition and analysis workflows have hindered wider adoption by the screening community. High-content, high-throughput tools for microscopy offer innovative and automated techniques for evaluating this complex biology. After the 3D cell culture is grown or transferred to an imaging microplate, cells can be treated with compounds and stained with the marker of choice. For acquisition, the ImageXpress® automated imaging systems and our cellular imaging and analysis software provide the capability to screen and analyze these models within a single, integrated interface to dramatically reduce time to discovery.

Steps used for a typical 3D cell culture assay:


3D cell culture imaging and analysis techniques

Our imaging technology continues to evolve for easier implementation of more physiologically-relevant models. Learn more about how our technology can help scale-up 3D cell culture from small-scale assays to high-throughput screening.



Solution for 3D Acquisition


Acquire images for 3D assays

In recent years, there have been significant advances in automated microscopy and imaging for delivering more predictive, physiologically-relevant measurements for drug discovery and environmental toxicity applications. Implementation of more complex assays and 3D models requires high resolution to capture publication-quality images and data.

Confocal imaging provides a high signal-to-noise ratio by reducing out-of-focus light for crisper images to yield more cellular detail even within a thick sample. Watch the video to learn about high-throughput techniques on how to acquire images in 3D using the ImageXpress Micro Confocal system.

Watch Video  


Solution for 3D Images


Analyze images for 3D applications

There are different types of image sets that may be generated even though your sample is 3D. A single plane may be sufficient or multiple planes may be acquired. When multiple planes are acquired, it’s useful to be able to analyze them individually or to analyze them after combining them into a single 2D projection. Ultimately, analysis of the entire 3D image may be the goal of the experiment. Watch the video to learn more about techniques for analyzing images from multiple z-planes as a 3D volume using MetaXpress software.

Watch Video    


Advantages of 3D Cell Models

Three-dimensional (3D) cell models are more physiologically relevant than two-dimensional cell cultures, and they more closely represent the tissue microenvironments, cell-to-cell interactions, and biological processes that occur in vivo. Now you can generate more predictive data using high-content imaging (HCI) systems like the ImageXpress system. By integrating both high-content image acquisition and analysis software like MetaXpress® software with the 3D Analysis Module, the ImageXpress system can be used for in-depth analysis, visualization, and assessment of 3D structures. This single interface will enable you to meet 3D acquisition and analysis challenges without compromise to throughput or data quality, giving you confidence in your discoveries.


Learn more about 3D cell model application and assays:

  • 3D Cancer Cell Research

    3D Cancer Cell Research

    Cancer involves changes which enable cells to grow and divide without respect to normal limits, to invade and destroy adjacent tissues, and ultimately to metastasize to distant sites in the body. Cancer spheroids mimic tumor behavior far more effectively than standard 2D cell cultures. Such 3D spheroid models are being successfully used in screening environments for identifying potential cancer therapeutics.

    Cancer researchers need tools that enable them to more easily study the complex and often poorly understood interactions between cancerous cells and their environment, and to identify points of therapeutic intervention.

    Learn More 

    3D neural spheroids

    3D neural spheroids

    To accelerate the development of effective and safe drugs, there is an increasing need for more complex, biologically relevant, and predictive cell-based assays for drug discovery and toxicology screening.

    We show that functional and morphological assays using 3D neural spheroids formed with human iPSC-derived cells can be used for evaluation of drug candidates and neurotoxicity assessment.

  • Cardiotoxicity


    Cardiotoxicity or cardiac toxicity is a term used to define chemicals that are toxic to the heart, causing muscle damage or heart electrophysiology dysfunction. The assessment of cardiotoxicity is important in the early stages of drug discovery to eliminate potentially toxic compounds from further development. Therefore, there is a growing need for highly predictive in vitro cardiotoxicity assays that use biologically relevant 3D cell-based models and are suitable for high-throughput screening.

    Learn More 

    Cells in Extracellular Matrices


    One common way of culturing cells in three dimensional space is through the use of extracellular matrix-based hydrogels, such as Matrigel. Cells are grown in an extracellular matrix (ECM) to mimic an in vivo environment. Differences between Matrigel and 2D cell cultures can be readily seen by their different cell morphologies, cell polarity, and/or gene expression. Hydrogels can also enable studies on cell migration and 3D structure formation, such as endothelial cell tube formation in angiogenesis studies.

  • Disease modeling using novel flowchip system

    Disease modeling using novel flowchip system

    Disease Modeling with 3D Cell-Based Assays Using a Novel Flowchip System and High-Content Imaging

    There is an increasing interest in using three-dimensional (3D) cell structures for modeling tumors, organs, and tissue to accelerate translational research. We describe here a novel automated organoid assay system (the Pu·MA System) combined with microfluidic-based flowchips that can facilitate 3D cell-based assays.

    Go to article

    Organoid Innovation Center

    Organoid Innovation Center

    The new Organoid Innovation Center at Molecular Devices combines cutting-edge technologies with novel 3D biology methods to address key challenges of scaling complex 3D biology.

    The collaborative space brings customers and researchers into the lab to test automated workflows for organoid culturing and screening, with guidance from in-house scientists.

    Learn more 

  • Organoid technology / Organ-on-a-Chip

    Organ on a Chip Assays

    Organoid technology, like organ-on-a-chip emulates organ physiology though co-culture of cells in a supportive 3D matrix and use of microfluidic channels to perfuse nutrients or compounds over the resulting cellular structures. It is rapidly gaining popularity as a biologically relevant screening model for new drugs and toxicity.

    Organoid vs. Spheroid

    Spheroid and Organoid

    The terms spheroid and organoid are often used interchangeably. Both cell models are 3D multi-cellular structures that better mimic the in vivo environment as compared to their 2D cell counterparts. Organoids typically consist of a co-culture of cells which demonstrate a higher order of self-assembly to allow for an even better representation of complex in vivo cell responses and interactions. In contrast, spheroids have a more simplified 3D structure making them amenable to high reproducibility and scaling. Functionally, both model types are used as better in vitro models for capturing more physiologically relevant data.

    Watch Video  

  • Organoids


    Organoids are three-dimensional (3D) multi-cellular microtissues that are designed to closely mimic the complex structure and functionality of human organs. 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.

    Learn More 



    Spheroids are multi-cellular 3D structures that mimic in vivo cell responses and interactions. They can be highly reproducible and to be scaled for high-content screening. Compared with adherent cells grown in 2D monolayers, 3D growth conditions are believed to more closely reflect the natural environment of cancer cells. Acquiring measurements from these larger structures involve acquiring images from different depths (z-planes) within the body of the spheroid and analyzing them in 3D, or collapsing the images into a single 2D stack before analysis.

    Learn More 

  • Stem Cells

    3D Stem Cells

    Stem cells are an accessible source of primary-like cells which can be used in longer-term studies to measure targeted responses in specific cell types and 3D tissues. Complex assays and 3D induced pluripotent stem cell (iPSC)-derived cell models better represent tissue biology and cell interactions which make them more relevant for many toxicity and drug screening assays.



    Toxicology is the study of adverse effects of natural or man-made chemicals on living organism. It is a growing concern in our world today as we are exposed to more and more chemicals, both in our environment and in the products we use.

    Read More 

  • Whole Organisms

    Zebrafish -Whole Organisms

    Model systems such as Drosophila, C. elegans, and zebrafish offer relevance to human biology, with the added advantage of genetic and whole-organism phenotypic screening. Zebrafish screening has gained favor as an alternative to mammalian screening due to reduced cost and space requirements, short generation times, and ease of genetic manipulation. Our zebrafish studies use the ImageXpress Micro System and MetaXpress software to set up and run automated imaging screens to monitor inhibition of angiogenesis, quantitate gene knockdown, and measure ototoxicity and neurotoxicity.

Latest Resources

3D cell imaging and analysis workflow

The workflow of a 3D cell culture protocol is dependent on the type of cell model being cultured. The 3D cell imaging and analysis workflow depicted here is a generalized example of 3D spheroids in a 96- or 384-well flat or round-bottom microplate. Each step breaks down the process from cell culture to image analysis and highlights the instrumentation and tools needed to conduct the cell culture protocol including an automated high-content imaging system and cellular imaging and analysis software.


3D Cell Imaging & Analysis Workflow
Culture Spheroids


3D cell can be cultured directly in a ULA, round bottom plate to develop the typical spherical shape of a spheroid. Cells can also be grown in alternate labware like a petri dish, flask, or other microplate before transferring to an imaging microplate.

96 or 384 Well Plates

96- or 384- Well Plates

Most 3D cell cultures for high-content, high-throughput screening are run using 96- or 384-well microplates. ULA, U-shaped/round bottom microplates enable the spherical shape to form spheroids or spheroids can also be cultured in a flat-bottom microplate using hydrogels to simulate extracellular matrices (ECM). Optimized spheroid cell culture protocols for the ImageXpress® Micro Confocal High-Content Imaging System can use ULA, U-bottom 96- and 384-well plates with a one-step staining procedure that reduces assay time and minimizes variability.


Shopping for reagents?

Check out our
EarlyTox Cell Viability Assay Kits

Treat with compounds


After spheroid formation, compounds are added into the wells, and then incubated for one to several days, depending on the mechanism being studied. Generally, shorter durations are used to study apoptosis and longer durations for multi-parameter cytotoxicity studies. For drug treatments requiring a longer duration, compounds are refreshed periodically during incubation. The concentration of compounds and the incubation period is dependent on the desired protocol.

Stain for Markers


After compound treatment is completed, stains are added directly to the media. Stains that require no washing can be used to avoid disturbing spheroids, but spheroids can be carefully washed even using automation, if necessary.

96 or 384 Well Plates

Assay Kits

Easy-to-use, robust assay kits for life science research, drug discovery and development, and bioassays. Our assay kits are optimized for use on our instruments. Screen more compounds earlier in drug discovery and enable characterization of a full concentration-response profile of test compounds for cell viability, cell proliferation, axiogensis, and more.

Shop assay kits  

Imaging System

High-content Imaging System

The ImageXpress Micro Confocal system is a unique confocal imaging solution capable of imaging more than a million wells a week. MetaXpress software enables acquisition of 3D z-stacks. Z-plane images can be saved individually or collapsed into a single 2D projection image using a mathematical algorithm.

View product  

Acquire 3D Cell Image


Images within the body of the spheroid can be captured individually or as a z-stack (multiple images taken at differing depths) using specialized imaging equipment.



Use cellular imaging analysis software to run quantitative analysis of the cell images to assess and monitor how cells express against different markers and to quantify biological readouts.

High-content Imaging Software

High-content Imaging Software

MetaXpress High-Content Image Acquisition and Analysis Software is a multi-level analysis tool for a wide range of 2D and 3D applications optimized for the ImageXpress Micro Confocal system. The integrated acquisition and analysis application modules for 3D cell models simplify high-throughput quantification of 3D structures with volume, intensity, and distance measurements.

View product  



Resources for 3D Cells

Videos & Webinars

Automate your 3D biology high-throughput workflows

Leverage automated, end-to-end workflows to enable complex organoid assays

Small Lab, Big Challenges - How the Right Reader can Impact Productivity

Level Up your 3D Cell Culture: From Research to High-Throughput

Enhance high-content 3D biology imaging with automated sample preparation

Enhancing 3D Disease Models: Automated, High-Throughput, Phenotypic Screening with Organ-on-a-Chip

Automating culture and high-content imaging of 3D organoids for in vitro assessment of compound effects

Getting Started with 3D Human Tissue Models and Imaging

ISSCR 2021 Innovation Showcase: Automated Culture and High-Content Imaging of 3D Lung and Cardiac

Organoid Innovation Center Walkthrough

Organoid Innovation Center Walkthrough

Capturing the complexity of 3D biology: Organoids for disease modelling and toxicity research

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

3D cell culture, tissue clearing, & high-content imaging in the quest of effective solutions to NAFLD

Transitioning high-content assays to 3D: Scientific opportunities and imaging challenges

Implementing 3D Neural Spheroids in Drug Discovery

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

Solutions for High-Throughput 3D Acquisition using High-Content Imaging

Solutions for High-Throughput 3D Acquisition using High-Content Imaging

Solutions for High-Content Analysis of 3D Images

Solutions for High-Content Analysis of 3D Images

Zebrafish Actin mask w FITC vessel mask

Zebrafish Actin mask w FITC vessel mask

Angiogenesis Research: High-Content Imaging Systems Help Unlock the Full Potential of 3D Tissue Models

Angiogenesis Research: High-Content Imaging Systems Help Unlock the Full Potential of 3D Tissue Models

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

Related Products And Services

The ImageXpress automated imaging systems are the ultimate combination of speed, sensitivity, and flexibility in a turnkey solution for your 2D or 3D applications and can accommodate different types of slide and plate formats. Our ImageXpress imaging systems allow superior visualization of thick samples by minimizing background fluorescence and increasing sharpness, resulting in accurate image segmentation. Increase the biological relevance of your screening assays with a seamless integration between acquisition and analysis of cells in a 3D space to yield volume, intensity, and distance measurements.