Comparison between different buffers and pressure-cooker times. CD20 (green) and CD30 (red) monoclonal antibodies have been used for the double immunofluorescence staining in a case of ALCL in combination with the break-apart ALK FISH probe. Results obtained with 2 min EDTA (a) show good immunofluorescence and FISH results. In some of the CD30+ cells (arrowheads), the FISH probe detected the rearrangement of the ALK gene, seen as separate red and green signals. Using 4 min citrate buffer (b) bad tissue preservation and the weak or absent immunofluorescence and FISH staining is observed.