An assessment on the use of tissue clear® versus xylene in deparaffinizing wax containing specimens for electron microscopy

Abstract

Electron microscopy plays an important role in diagnostic histopathology. When this investigation is anticipated, extra tissue is submitted directly for electron microscopy. However, often it is decided only later in a problematic case to perform this investigation and then the only tissue available is embedded in the routine laboratory’s paraffin block. This tissue has to be retrieved from the wax and this entails using a clearing agent before the rest of the electron microscopy method can be implemented. Traditionally xylene is the agent that is used but has the disadvantage of being extremely toxic.

This study compared the morphological effects of a relatively new and non toxic clearing agent, Tissueclear®, with that of xylene. Exposure of tissue to clearing agents for 30 minutes and overnight was performed to assess whether Tissueclear® gave better results in the long term than xylene, in the hope that the laboratory turn around time could be improved and the amount of toxic reagents used in the EM laboratory will be reduced. A second part of the study involved a questionnaire submitted to laboratory staff to assess their knowledge of xylene toxicity.

Of the 325 cases submitted for electron microscopy at Universitas Hospital between January 2004 and July 2005, 140 of these had to be retrieved from paraffin wax. Four specimens were prepared from each case. Two were processed in xylene for 30 minutes and overnight and two in Tissueclear® for 30 minutes and overnight. The specimens were evaluated for consistency and resin compaction as well as ultrastructural preservation of the cell membrane, cytoplasmic content and extracellular material.

The results showed that Tissueclear® and xylene gave comparable results after 30 minutes and that Tissueclear® was superior after overnight processing.

This meant that a specimen submitted for electron microscopy would be processed immediately without waiting for the following morning as was the case with xylene and that the processing time for such a specimen had been shortened from 3 to 2 days. It also meant that the laboratory staff was exposed to one less toxic reagent.

The results on the questionnaire showed that there were large areas of ignorance regarding toxicity as well as appropriate safety procedures that need to be followed in the laboratory. It is hoped that this study will improve awareness in this regard and encourage the use of other newer less toxic reagents.