On colouring epon-embedded tissue sections with Sudan black B or Nile blue A for light microscopy

Abstract

ABSTRACT Sections of osmium-fixed tissues embedded in epon 812 colour with either Sudan black B or Nile blue A solutions to reveal a variety of detail by direct microscopy with normal apochromatic or semi-apochromatic objectives. The clarity of the coloration gives a picture fully comparable to that seen by phase-contrast microscopy. The plastic is not removed. 1μ sections or thinner sections down to green or gold, are mounted on clean glass slides by drying down from 20% acetone/water after flattening on a hot plate. Colouring is carried out at room temperature in Sudan black B (saturated solution in 70% alcohol) for 1 to 2 h. The result is a reversed or negative effect, for the epon plastic takes the stain avidly, but dense elements of the tissue do not, and appear white against a blue background of stained plastic. Lipid droplets retain a capacity to colour, becoming dark blue to blue-black. Nile blue sulphate (1% aqueous solution) colours thin sections of tissue in 1 to 2 h at 60° C, acting apparently as a basic dye on most cell elements, and also colours lipid droplets dark blue. After both techniques the sections are mounted in Farrants’s medium.