Reduction of autofluorescence in the postnatal and adult spiral ganglion by quenching with CuSO₄ and Sudan Black B

Introduction  The phenomenon of autofluorescence in histologic sections of the cochlea is primarily due to natural fluorophors such as lipofuscins and elastin. This autofluorescence represents a considerable interfering signal during microscopic detection of fluorescent markers. The aim of this work was to quantify autofluorescence in the rat spiral ganglion at different ages and to evaluate different experimental approaches to reduce this interfering signal.

Material and Methods  In cryosections of the neonatal (P0), postnatal (P4) and adult (7 months) rat cochlea, we used gray-scale analysis to quantify autofluorescence in the spiral ganglion. Specimen were treated with CuSO4 and Sudan Black B at different concentrations and incubation times and the effects were measured.

Results  Prominent autofluorescence was detected at all three ages analyzed. While CuSO4 led to a significant reduction of autofluorescence only in the postnatal (P4) spiral ganglion, Sudan Black B significantly decreased the interfering signal at all ages. The optimal reduction in autofluorescence was obtained by combined treatment with both substances.

Conclusion  Autofluorescence of the spiral ganglion leads to a severe disturbance of the microscopic analysis of fluorescent markers in neo-/postnatal as well as in adult animals. Treatment of the tissue with Sudan Black B and CuSO4 allows a significant reduction of the interfering signal by quenching the autofluorescence. However, the age of the animals has to be considered when choosing the optimal concentration and exposure time.

Publication History

Article published online:
24 May 2022