Elimination of Autofluorescence in Archival Formaldehyde-Fixed, Paraffin-Embedded Bone Marrow Biopsies

Abstract

Context.— High levels of autofluorescence in bone marrow tissue constitute a major obstacle to immunofluorescence analysis of bone marrow biopsies. Objective.— To present a simple, efficient method to eliminate autofluorescence in bone marrow biopsies. Design.— Autofluorescence of paraffin bone marrow tissues was examined in different hematologic disorders with confocal laser scanning microscopy. Strong autofluorescence was observed in primary myelofibrosis and acute leukemia with reticulin myelofibrosis in 488-nm and 561-nm channels. To eliminate autofluorescence, AutoFluo Quencher was used on bone marrow sections with different incubation times. The effects of AutoFluo Quencher on immunofluorescence analysis of bone marrow biopsies was tested using antibodies tagged with different fluorophores. Results.— AutoFluo Quencher thoroughly eliminated the strong autofluorescence of bone marrow but did not decrease the intensity of fluorophores, leaving the specific signals of target proteins clearly visible. Conclusions.— This study presents a simple, efficient method to eliminate autofluorescence in bone marrow paraffin tissue, and it opens the way to better results in the immunofluorescence analysis of bone marrow biopsies.