Immunofluorescent visualization of CD11b/c and ionotropic glutamate receptor subunit GluR4 in neonatal rat brain microglia. The integrin CD11b/c and the ionotropic glutamate receptor subunit GluR4 were visualized using the dual-labeling indirect immunofluorescent technique (See Materials and Methods). (Top left) bright field, phase contrast view of microglia incubated in the absence of the primary antibodies against GluR4 and CD11b/c. (Top right) same microglia viewed using FITC filter configuration. Note vesicular structures exhibiting intense autofluorescence, but little or no fluorescence over most of the microglia cell bodies or the elongated processes. (Middle left) bright field, phase contrast view of microglia labeled with anti-glutamate receptor GluR4 subunit and the leukocyte marker CD11b/c. (Middle right)(green) same field viewed using the FITC filter configuration. Microglia not only contain fluorescent vesicles, but exhibit a prominent speckled pattern of fluorescence over the cell bodies and the elongated processes. Absence of speckled pattern of labeling in cells incubated without primary GluR4 antibody (compare with top right photo) suggests the speckled pattern represents specific labeling of GluR4 subunit. (Lower left) same microglia exhibited intense labeling of CD11b/c (red), which confirms microglia are of leukocytic origin. (Bottom right) GluR4 and CD11b/c superimposed labeling (yellow). Approximate magnification of the original was × 525.