Rapid Freezing of Small Biopsies and Standard for Cryosectioning and X-Ray Microanalysis

Abstract

A method for clamp-freezing of small biopsies for ultrathin cryosectioning is described. A formvar film is used to support the specimen before freezing with a pair of pliers, precooled in liquid nitrogen. This method makes it possible to align the piece of tissue close to a drop of standard solution and freeze the two simultaneously. After clamping, the flat, frozen sample contains both the piece of tissue and the standard in one solid block, which may thus be sectioned as a whole. The method has been used in the surgical theater, yielding good results with needle biopsies of the prostate and cystoscopically obtained biopsies of bladder tumors. The time lapse between excision and freezing may be less than 30 seconds, which represents a great improvement compared with the far more time-consuming mincing of tissue and mounting of the pieces on metal pins. From the frozen specimen, a large number of sections with good morphologic characteristics may be produced. The method was evaluated with the use of rat liver and lactating mammary tissue. Directly frozen tissue with intact blood supply was compared with biopsies prepared as described. Redistribution of Ca could not be demonstrated in either specimen type. However, the results indicate that the monovalent ions are far more mobile, and a significant increase of the Na:K ratios was observed in the biopsies compared with directly frozen tissue.