Some Experiments with Chromium Compounds as Fixers for Electron Microscopy

In the course of experimentation with various tissue fixations other than Os04 it was noticed that mixtures containing chromium compounds showed considerable promise. Therefore the fixation characteristics of three such compounds, CrC13, K2Cr~07 and Cr08 (the anhydride of H2Cr04), were systematically investigated. Although the action of all three was essentially similar, CrO~ proved to be the m o s t satisfactory and was used exclusively in subsequent experiments. Its concentration was varied from 1 to 6 per cent. It was tried in combination with formaldehyde, the Concentration of which was varied from 2 to 32 per cent. Sodium chloride, in concentrations never exceeding 3.4 per cent, was used t o control tonicity in many experiments. A number of standard buffers were mixed with the solutions without noticeable profit. The fluid finally adopted as the most satisfactory was an aqueous solution containing 3 per cent chromic oxide (CrOa), 4 per cent formaldehyde (I-ICHO) and 0.85 per cent sodium chloride (NaC1). The ptI of this mixture was about 3.2. The fixation time was varied from 5~ to 17 hours, but after several hours fixation * Aided bygrants from the Public Health Service (H-1663) and The Lilly Research Laboratories. ~t Public Health Training Fellow (HTS-sill). the tissue resembled a metal-shadowed preparation, (apparently due t o gradual deposition of chromium) and so it was decided that a short fixation of k~ :to ~ hour was best. Washing for ~ hour in distilled water immediately after:fixation produced "cleaner" preparations. Since cells rich in endoplasmi¢ reticulum such as those of the exocrine pancreas responded most favorably t o this type of fixation, these were adopted as test ob-jeets in developing the procedures, Occa. sionally the somewhat less basophilic parotid cells were also used. After methacrylate embedding the tissue was light yellow or brown with a darker center and the dark areas deep in the tissue ustmlly represented areas of better fixation. The tissue was brittle and hard, but with care could be sectioned in silver ribbons with a Porter-Blum micro-tome. Despite the common presence of fine chatter good sectiOns Were obtained from almost every block. The general appearance of pancreatic acinar cells fixed by this method is: fllns. trated in Figs. 1 to 4. The spaces within the endoplasmic reticulum appear less dense and alternate throughout most of the cytoplasm with darker bands representing : the cytoplasmic matrix. The apparently linear organization of: the cytoplasm is …