Phosphotyrosine antibodies specifically label ameboid microglia in vitro and ramified microglia in vivo

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Abstract
Using an affinity‐purified, polyclonal antibody to phosphotyrosine (Wang: Molecular and Cellular Biology 5:3640–3643, 1985) we have previously demonstrated that phosphotyrosine immunoreactivity is restricted to a population of multipolar GFAP‐negative neuroglia in adult rat brain (Tillotson and Wood: Journal of Comparative Neurology 282:133–141, 1989) and retina (Tillotson and Wood: Journal of Cell Biology 107:724a, 1988). In this study, we show that the phosphotyrosine‐immunoreactive cells are microglia. This conclusion is supported by numerous morphological and ultrastructural similarities between the phosphotyrosine‐immunoreactive cells and microglia. Furthermore, phosphotyrosine co‐localizes with the microglial‐specific B4 isolectin of Bandeiraea simplifolia‐1 lectin. Phosphotyrosine antibodies also stain ameboid microglia in primary cultures of neonatal rat brain. In addition, after 7 days in vitro, microglia are the only phosphotyrosine‐immunoreactive element in the cultures. This temporal pattern of staining in vitro mimics the developmental progression of phosphotyrosine immunoreactivity in situ, in which a variety of structures stain during postnatal neural development (Tillotson and Wood: Journal of Comparative Neurology 282:133–141, 1989), but only microglia stain in mature brain. The significance of phosphotyrosine‐containing proteins potentially expressed in a microglial‐specific manner is discussed.