Protocol to generate microglia-containing cerebral organoids to model HIV neuroinflammation

  • STAR Protoc. 2026 Jun 19;7(2):104604. doi: 10.1016/j.xpro.2026.104604.
Janet P Zayas  1 ,  David Gagliardi  1 ,  Stephanie Gambut  1 ,  Sam Acors  2 ,  Nathalia Almeida  2 ,  Michael H Malim  2 ,  Luis Apolonia  3 ,  Lena Al-Harthi  1 ,  Srinivas D Narasipura  4 ,  João I Mamede  5
Affiliations
  • 1. Department of Microbial Pathogens and Immunity, Rush University Medical Center, Chicago, IL 60612, USA.
  • 2. Department of Infectious Diseases, School of Immunology and Microbial Sciences, King's College London, London SE1 9RT, England.
  • 3. Department of Infectious Diseases, School of Immunology and Microbial Sciences, King's College London, London SE1 9RT, England; Centre for Gene Therapy & Regenerative Medicine, School of Basic & Medical Biosciences, King's College London, London SE1 9RT, England.
  • 4. Department of Microbial Pathogens and Immunity, Rush University Medical Center, Chicago, IL 60612, USA. Electronic address: [email protected].
  • 5. Department of Microbial Pathogens and Immunity, Rush University Medical Center, Chicago, IL 60612, USA. Electronic address: [email protected].
PMID: 42224077 DOI: 10.1016/j.xpro.2026.104604
Abstract

Cerebral organoids (COs) are valuable for studying neurodegenerative diseases and pathogens, but their limited microglia pose challenges. Here, we present a protocol to generate organoids with physiologically relevant microglia (CO-iMs). We describe steps for differentiating induced pluripotent stem cells (iPSCs) into mesoderm, seeding embryoid bodies (EBs), and harvesting hematopoietic progenitor cells (HPCs). We detail procedures for co-culturing HPCs and iPSCs to generate CO-iMs, followed by their Infection and treatment. The model provides a platform to study viral infections and neuroinflammation. For complete details on the use and execution of this protocol, please refer to Narasipura et al.1.

Keywords
Biotechnology and bioengineering; Microbiology; Organoids; Stem Cells.
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