Microglia applications & functional biology

What are the main research applications for human iPSC-derived microglia today?

iPSC‑derived microglia offer a reproducible and scalable platform for disease modeling and compound screening. They can be used in disease-agnostic assays by evaluating the effect of compounds on cell activation, for example, by measuring cytokine release. Microglia function can also be assessed more specifically by investigating phagocytic uptake of labeled bait, such as beta-amyloid, alpha-synuclein, dead neurons, and many more.

Because iPSC technology allows the use of patient‑derived, gene‑edited, or control lines, researchers can explore how microglial function differs between health and disease. Additionally, iPSC platforms support patient lines alongside CRISPR-engineered isogenic controls, and researchers can isolate mutation‑driven differences in microglial behaviour. When combined with neurons in co‑cultures, microglia also enable the investigation of cell‑to‑cell interactions and how microglial activation can impact neuronal health and function.

What is microglia’s role in ophthalmology research?

Microglia, the resident immune cells of the retina, play a crucial role in the pathogenesis of dry age-related macular degeneration (dry AMD). During AMD progression, microglia become activated in response to oxidative stress, the release of pro-inflammatory cytokines, and drusen deposits in the retinal pigment epithelium (RPE).

The role of microglia activation in AMD remains unclear. While it may be initially protective by contributing to debris clearance, chronic activation leads to a proinflammatory state characterized by the release of proinflammatory cytokines and complement factors, which could contribute to photoreceptor degeneration and geographic atrophy, a hallmark of late-stage dry AMD. Understanding microglial involvement in dry AMD offers novel potential therapeutic targets to modulate their activity and reduce neuroinflammation-associated retinal degeneration. Learn more in our ophthalmology eBook: https://axolbio.com/publications/physiologically-relevant-in-vitro-retinal-models-for-ophthalmology-drug-discovery-and-safety-testing/

How do researchers use microglia models to evaluate neurotoxic vs. neuroprotective drug effects?

Microglia can be added to neuron-astrocyte co‑cultures to investigate the interplay between these cells. Microglia can be activated and the effect on neuronal health and function assessed. Once activated, their effects on neuronal firing and network behavior can be measured using multi‑electrode array recordings or calcium imaging platforms. These tools allow researchers to evaluate how firing dynamics change in the presence of microglia, with or without stimulatory or stress conditions. Patient‑derived or gene‑edited microglia can also be used to study disease‑specific effects on otherwise healthy neurons.

How important is incorporating disease-relevant genetics (APOE4, TREM2) into microglia models? How do iPSC systems support this?

Using iPSC‑derived microglia carrying mutations such as APOE4 or TREM2 allows assessing phenotypic differences in a physiologically relevant system. These models enable comparison of healthy and disease backgrounds, support mechanistic studies, and facilitate large‑scale compound screening and lead candidate selection. Incorporating these lines into more complex co‑culture systems allows investigation into how disease‑relevant microglial states influence other neuronal cell types.

Microglia maturation, ontogeny & phenotypic diversity

Would more controlled maturation and stable phenotypes from iPSC-derived microglia improve functional assay consistency?

Yes. At Axol, our iPSC‑derived microglia undergo extensive characterization and batch‑to‑batch testing to ensure consistent morphology, marker expression, and functional responses. This is essential for reliable in vitro research, disease modeling, and screening workflows. The result is a physiologically relevant, scalable microglia model suitable for diverse applications. See our microglia characterization: https://axolbio.com/publications/generation-and-characterization-of-cryopreserved-axocells-human-ipsc-derived-microglia-2/

What are the challenges in faithfully recapitulating in vivo microglial states?

Effectively recapitulating in vivo microglial states (resting, activated, or transitioning) is challenging because each available microglial model captures different aspects of native biology. Rodent microglia, primary human microglia, immortalized cell lines such as HMC3, and iPSC‑derived microglia each contribute valuable information depending on the question being asked.

Modeling in vivo microglial states comes with several challenges: translatability, scalability, and availability. Species differences can make it difficult to translate findings directly into human biology, while access to primary human microglia is limited, making them an unsuitable and non-scalable resource for high‑throughput screening. Although immortalized cell lines provide larger cell numbers, they may be limited in physiological relevance and disease-specific modeling.

iPSC-derived microglia help address many of these limitations by demonstrating key functional characteristics of in vivo microglia. They can be driven to an activated state, show cell plasticity, phagocytic uptake of multiple baits, changes in morphology, and express key cell-specific markers (Iba1, TMEM119, PU.1, CX3CR1). iPSCs offer a balance of reproducibility, scalability, and physiological relevance, and support studies across genetically diverse backgrounds.

Microglia co‑culture & tri‑culture systems

What co‑culture systems are commonly used with microglia?

Microglia can be integrated into a range of co‑culture systems depending on the disease area. In ALS modeling, they are combined with motor neurons and astrocytes. In neuroinflammatory or Alzheimer’s research, they are added to networks containing excitatory cortical neurons, inhibitory interneurons, and astrocytes. Combining striatal neurons, astrocytes, and microglia in vitro enables physiologically relevant co‑culture models that replicate HD phenotypes and reveal region‑specific and cell‑cell drivers of neurodegeneration. For dry AMD, they are co‑cultured with RPE cells to explore inflammatory interactions in the retina.

How can microglia-neuron co-cultures improve our understanding of disease mechanisms?

Multi‑cell systems allow researchers to observe how microglial activation influences neuronal function. By stimulating microglia within these cultures, it is possible to evaluate effects on neurons or other glial cells. Using disease‑derived microglia in the same environment as healthy neurons can help observe how pathological microglial states affect network activity, synaptic health, and inflammatory balance.

Are tri-cultures being used for AD/ALS/HD research, and does using matched iPSC-derived cell types reduce variability?

Yes, tri-cultures are actively being developed at Axol across several disease areas. Key considerations include media formulations to support all cell types, compatible cell densities, and determining the appropriate timing for microglial introduction.

Using isogenic cell types allows researchers to build a more controlled and physiologically relevant system, making it easier to interpret how each cell type contributes to the model. Introducing non‑isogenic cells can also be valuable, as it helps reveal how a diseased cell influences other cells within the system and contributes to dysfunction. Learn more: https://axolbio.com/publications/validation-of-a-cortical-tri-culture-axomodel-for-in-vitro-compound-screening-a-blinded-compound-study/

Microglia in organoid & 3D systems

Why are microglia important for brain organoid maturation and disease modeling?

Neuroinflammation is a key component of many neurodegenerative conditions, and is often a common factor across them. Dysfunction in microglia, including decreased phagocytosis, increased inflammatory signaling, and chemotaxis, contributed to disease progression and can drive neurotoxicity and dysfunction in other glial cells. Microglia also play essential roles in maintaining homeostasis within the CNS, supporting a physiological environment that promotes neuronal maturation and firing. Introducing microglia into organoid systems is therefore an important step toward recapitulating a physiological environment for the cells and driving the maturity and functionality of the model. If microglia are absent, organoids may remain transcriptionally immature and lack fully developed physiological network activity.

How are microglia incorporated into cerebral organoids or assembloids? Can Axol microglia be integrated into organoids?

Microglia can be added to cerebral organoids or assembloids at defined developmental stages, where they migrate, integrate, and begin interacting with surrounding neural cells. High‑quality, stable microglia are important for successful incorporation, as they support maturation, immune responsiveness, and long‑term function within the 3D environment. Researchers have been successfully integrating axoCells in organoid workflows, and our experience in building complex 2D models helps guide their application in 3D systems.

High‑throughput & screening applications

Can iPSC-derived microglia be used in high-throughput drug screening workflows?

Yes. In collaboration with Sygnature Discovery, we demonstrated the utility of our iPSC‑derived microglia in automated, miniaturized screening systems, including 384‑well platforms and compound libraries comprising more than 1000 compounds. Learn more about this collaboration: https://library.axolbio.com/document/sygnature-discovery-poster-automation-of-ipsc-workflows/

What challenges exist when scaling microglia cultures for multi-well formats?

While a microglia differentiation run can produce ~200 million cells, vials are frozen at 1 million viable cells to ensure consistency and viability. As a result, high‑throughput formats require multiple vials, which can restrict larger screening efforts. As interest in larger screening formats grows, alternative cryopreservation densities and packaging options may further streamline these workflows.

Can high-throughput platforms identify compounds that modulate microglial activation?

Yes. High‑throughput systems can effectively measure functional endpoints such as cytokine release, phagocytosis, and imaging‑based morphological changes. Compounds can be tested in 384‑well formats, allowing robust, high‑quality data generation.

Future advancements & emerging technologies

What next-generation approaches are shaping the future of microglia research?

The role of microglia in neurodegeneration has been recognized for many years, and being able to study patient‑derived microglia across different diseases now offers valuable insight into disease modeling and drug discovery. Next‑generation approaches in microglia research are centered on building physiologically relevant and integrated systems. The next step is to examine these cells within multi‑cell 2D and 3D systems to better understand how they interact with other neural cell types, how dysregulated inflammatory signals drive neuronal dysfunction, and whether these effects can be reversed or improved. Techniques such as single‑cell sequencing are also refining our understanding of the diverse microglial states present in disease. Together, these approaches are enabling more precise models of disease mechanisms and advancing microglia‑focused therapeutic development.

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