Interference microscopy delineates cellular proliferations on flat mounted internal limiting membrane specimens.

摘要

Aim: To demonstrate that interference microscopy of flatmounted internal limiting membrane specimens clearlydelineates cellular proliferations at the vitreomacularinterface.Methods: ILM specimens harvested during vitrectomywere fixed in glutaraldehyde 0.05% and paraformaldehyde2% for 24 h (pH 7.4). In addition to interferencemicroscopy, immunocytochemistry using antibodiesagainst glial fibrillar acidic protein (GFAP) and neurofilament(NF) was performed. After washing in phosphatebufferedsaline 0.1 M, the specimens were flat-mountedon glass slides without sectioning, embedding or anyother technique of conventional light microscopy. A coverslide and 49,6-diamidino-2-phenylindole (DAPI) mediumwere added to stain the cell nuclei.Results: Interference microscopy clearly delineatescellular proliferations at the ILM. DAPI stained the cellnuclei. Areas of cellular proliferation can be easilydistinguished from ILM areas without cells.Immunocytochemistry can be performed without changingthe protocols used in conventional microscopy.Conclusion: Interference microscopy of flat mounted ILMspecimens gives new insights into the distribution ofcellular proliferations at the vitreomacular interface andallows for determination of the cell density at the ILM.Given that the entire ILM peeled is seen en face, thetechniques described offer a more reliable method toinvestigate the vitreoretinal interface in terms of cellulardistribution compared with conventional microscopy.