Confocal laser scanning microscopy of retinal rod outer segment intact disks: new labeling technique

Abstract

Vertebrate retinal rod outer segment disks house the proteins involved in the phototransduction cascade that converts light into neuronal signal. We develop a technique for the immunofluorescent labeling of osmotically intact isolated rod outer segment disks for confocal laser scanning microscopy imaging. Osmotically intact Ficoll-flotation isolated bovine disks are directly labeled with antibodies in solution. For the first time, osmotically intact single disks can be visualized. Thus, imaging of purified disks, based on advanced optical techniques, may serve as a powerful complement to other methods in studies on phototransduction. In fact, even though much is known about the rod outer segment photoresponse, some unanswered questions remain, particularly about ATP supply, light adaptation, and morphogenesis.