Enhanced visualization of weak colloidal iron signals with Bodian's protein silver for demonstration of perineuronal nets of proteoglycans in the central nervous system

The present study aimed for a clear visualization of faintly deposited coll oidal iron in tissue sections for light microscopy. Paraffin blocks contain ing paraformaldehyde-fixed brain tissue from healthy adult mice were cut in to sections 10-15 mum thick. After deparaffinization, the sections were sta ined with fine cationic iron colloid at a pH value of 1.0-1.5, and treated with a mixture of potassium ferrocyanide and hydrochloride for Prussian blu e reaction. Some sections were further treated with Bodian's protein silver after the Prussian blue reaction. This sensitized development of Prussian blue reaction with Bodian's protein silver more clearly visualized the fain tly deposited cationic colloidal irons than the demonstration by Prussian b lue reaction alone, and allowed an enhanced visualization of the perineuron al nets of sulfated proteoglycans in the brain. Thus, such fine perineurona l sulfated proteoglycans as those in the CA3 field of the hippocampus, whic h are weakly stained with cationic iron colloid acid usually overlooked by a demonstration with only a Prussian blue reaction, could be clearly visual ized with striking contrast by the sensitized development with Bodian's pro tein silver after the Prussian blue reaction. Preliminary hyaluronidase dig estion erased Bodian's protein silver development of perineuronal sulfated proteoglycans, Though some axonal fibers were also additionally stained wit h Bodian's protein silver itself, this sensitized development is useful to enhance such weak colloidal iron signals as are hardly detectable by only P russian blue reaction.