Alcian dyes in calcium chloride: a routine selective method to demonstrate mucins.

One of the simplest procedures to demonstrate mucinous substances (acid mucopolysaccharides) in routine histological preparations involves the use of Alcian dyes. Methods involving these dyes have largely. replaced the older, more empirical mucicarmine technique arid are faster and less complicated than the colloidal iron method of Hale,1 which suffers from the possibility of false-positive staining due to the presence of intrinsic iron in tissues.? The usefulness of the Alcian dye methods for diagnostic purposes has been extended by combination with other procedures. Mowry8 used Alcian blue-periodic-acid-Schiff method to demonstrate the acidic groups and the 1,2 glycols of mucopolysaccharides. Putt and Hukill'. found Alcian green-Verhoeff's elastic stain useful, for the histologic: diagnosis of lung carcinomas. Alcian blue 8GS, introduced by Steedmand it a water soluble, copper phthalocyanine dye, which is closely related to the alcohol soluble Mona §stral fast blue. Since Alcian blue 8GS is no longer commercially available, it has been replaced by Alcian blue 8GX (C.I. No. 74240), a purer and more soluble dye. The original reaction for demonstrating mucins was of necessity very short: if prolonged, other tissue elements took up the stain. This was presumably due .to both strong and weak acid groups in tissue, the former responsible for the fast reaction. In order to overcome this problem, Vailli oxidized sectioned tissues in chromic acid, which limited the staining reaction to the strong acidic groups of mucopolysaccharides. Mowry' and Lison8 further modified the staining procedure by acidifying the dye solution with acetic acid, which permitted a longer and more precise stain with minimal background coloration due to the fact that the low pH allowed only the utilization of strong acid esters. The slight nuclear or connective tissue staining that sometimes occurs with Alcian dyes can be overlaid and minimized with a counterstain. The purpose of this paper is to introduce a method of preparing Alcian dye solutions by the substitution of calcium chloride (CaCI2.2H2O) in place of acetic acid. This replacement results in a solution with a pH of 2.8 which is well within the limits of acidity of the solutions used by Mowry and Lison. The present selective use of Alcian dyes for mucin in a calcium salt solution was first observed inadvertently while carrying out Attwood's differential