Table 2.

Main mouse lines used to visualize microglia from early embryonic development to adulthood and aging Other proteins expressed by microglia but whose specificity is not confirmed include APOE, CLEC7A, ITGAX, and LPL.

Mouse line	Specificity	Labeled states	Expression patterns	Main applications	Reference
CX3CR1-GFP	macrophages including microglia	homeostatic conditions and disease associated, but downregulated in DAM, MGnD, dark microglia, and other pathological states.	visualization of microglial cell body and processes. fluorescence diffuses throughout the cytoplasm. bright enough for two-photon in vivo imaging. a limitation is that the heterozygous mice used for in vivo imaging are partially deficient in fractalkine signaling, with possible outcomes on the brain and behavior.²⁰⁶ The homozygous mice are knockout for CX3CR1 and used to study the outcomes of fractalkine receptor deficiency.	two-photon in vivo imaging or fluorescence analysis of microglial density, distribution, dynamics, interactions with other parenchymal elements, and categorization into morphological states. ultrastructural studies using staining against GFP.	Davalos et al.;⁵⁵ Nimmerjahn et al.;⁵⁶ Jung et al.;¹⁷⁸ Bisht et al.;¹⁸⁰ Tremblay et al.;¹⁸⁵ Paolicelli et al.²⁰⁷
Iba1-EGFP	macrophages including microglia	homeostatic conditions and disease associated. downregulated in some contexts (e.g., obesity and aging) and in some pathological states (e.g., DAM, dark microglia). used to study microglia in early embryonic and postnatal development. conserved across several species including human.	visualization of microglial cell body and processes. fluorescence diffuses throughout the cytoplasm. less bright than fluorescence in CX3CR1-GFP mice but generally sufficient for two-photon in vivo imaging of cell body and proximal processes. these mice are not partially deficient in IBA1 in their heterozygous state, which is a main advantage.	two-photon in vivo imaging or fluorescence analysis of microglial density, distribution, dynamics, interactions with other parenchymal elements, and categorization into morphological states. ultrastructural studies using staining against GFP.	Bisht et al.;¹⁸⁰ Wake et al.;¹⁸⁴ Hirasawa et al.²⁰⁸
Fms-EGFP or CSF1R-EGFP; CSF1R-FusionRed	Macrophages including microglia. CSF1R is expressed by most microglia.	Homeostatic conditions and disease-associated, but considered to be downregulated in DAM and other pathological states.	Fluorescence is less bright than in CX3CR1-GFP mice, and generally sufficient for two-photon in vivo imaging. It also allows for fluorescence-activated cell sorting and fluorescence imaging when combined with immunostaining. These mice are not partially deficient in CSF1R in their heterozygous state, which is a main advantage.	Fluorescence-activated cell sorting and fluorescence analysis of microglial density, distribution, dynamics, interactions with other parenchymal elements, and categorization into morphological states when combined with immunostaining.	Grabert et al.;³⁴ Sierra et al.;¹⁶² Sasmono et al.²⁰⁹
HEXB-TdTomato	largely overlaps with IBA1 staining but restricted to microglia. Does not label CAMs and other border-associated macrophage populations.	expression appears stable in homeostatic conditions and disease-associated states. The labeled microglia are also depleted by CSF1R inhibition.	visualization of microglial cell body and processes. fluorescence diffuses throughout the cytoplasm. bright enough for two-photon in vivo imaging. a limitation is that the heterozygous mice used for in vivo imaging are partially deficient in HEXB. However, their microglial gene expression patterns do not appear affected.	two-photon in vivo imaging or fluorescence analysis of microglial density, distribution, dynamics, interactions with other parenchymal elements, and categorization into morphological states.	Masuda et al.³⁸