Dr Burns and colleagues reply

A recent article by Burns et all referred to a silver amplification system that was crucial to the success of the demonstration, by in situ hybridisation, of specific sequences of DNA in the sex chromosomes. The silver amplification system was used to visualise complexes of diaminobenzidine (DAB), and uses the principles of a method established by ourselves-namely, that complexes of DAB bind gold salts,2 and that the avidity of DAB for gold salts (gold chloride in particular) is so great that it can be used to generate an insoluble sulphide on which the silver amplification procedure depends.34 We were surprised that as we were the first to have used these sequential steps to enhance DAB visualisation no mention was made of our work and that the reference given for the use of gold salts was to a brief letter,5 which, in fact, makes no mention of the use of gold. In our publications we have acknowledged the earlier work of Siegesmund et al who, in an x ray microanalysis study, showed the affinity of DAB for gold salts.6 We would not like your readers to believe that the application of the silver amplification procedure described by Burns et al is confined to in situ hybridisation technology. The method by which the DAB is generated is irrelevant: silver amplification based on the principles that we have described can be used for various applications, ranging from work on artificial materials such as nitrocellulose to tissue sections for the light and electron microscope.