TY  - JOUR
TI  - Protocol for Mouse Embryonic Aorta–Gonad–Mesonephros (AGM) Region Frozen Sectioning and Immunofluorescence
AU  - Jia, Yan 
AU  - Huo, Sixuan 
AU  - Li, Mei 
VL  - 16
IS  - 18
PY  - 2026
DA  - 2026/09/20
SP  - e5828
C1  - Bio-protocol 2026;16:e5828
DO  - 10.21769/BioProtoc.5828
UR  - https://doi.org/10.21769/BioProtoc.5828
AB  - The aorta–gonad–mesonephros (AGM) region is the site where hematopoietic stem cells (HSCs) first emerge during development, and is therefore widely used to study in vivo hematopoiesis and to discover novel regulatory mechanisms. The endothelial-to-hematopoietic transition (EHT) process can be directly observed via immunofluorescence on frozen sections of the AGM region. However, the mouse AGM region is extremely delicate and lies deep within the embryo, between the notochord and the somatic mesoderm. Here, we present a step-by-step protocol covering embryo collection, fixation, dehydration, and embedding with a defined orientation, followed by frozen sectioning, immunofluorescence staining, and confocal imaging. The protocol is highly reproducible and easy to follow and provides clear instructions on orienting the embryo and anatomically locating AGM. By filling a technical gap, the protocol can enable researchers to reliably study HSC emergence and EHT in the mouse embryonic AGM.
KW  - Aorta–gonad–mesonephros
KW  - Hematopoietic stem cells
KW  - Endothelial-to-hematopoietic transition
KW  - Cryosectioning
KW  - Immunofluorescence
JF  - Bio-protocol
SN  - 2331-8325
PB  - Bio-protocol LLC.
BIO101  - False