Utilization of Z-6040 organosilane as a coupling agent to improve the adhesion of epoxy resins to waxy biological tissues

Authors: Mowery, Joseph; Bauchan, Gary

Submitted to: Microscopy and Microanalysis
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 4/24/2020
Publication Date: 7/30/2020
Citation: Mowery, J.D., Bauchan, G.R. 2020. Utilization of Z-6040 organosilane as a coupling agent to improve the adhesion of epoxy resins to waxy biological tissues. Microscopy and Microanalysis. 26(Suppl 2):1352-1353. https://doi.org/10.1017/S143192762001781X.
DOI: https://doi.org/10.1017/S143192762001781X

Interpretive Summary: When preparing plants or insects for high resolution imaging using a transmission electron microscope, the specimens must first be incased in a hard resin and sliced into thin sections. A commonly encountered obstacle when embedding waxy biological tissue such as plants or insects is the tendency of the resin to detach from the sample during slicing. Here we report on the use of Z-6040 as an additive during the tissue processing stage. We examined the adhesion of the resin to eggs of the squash bug and leaves of sweet orange. Ultimately, samples embedded without Z-6040 were observed to have small gaps along the perimeter of the specimen cuticle where the resin had separated from the tissue. Samples which were primed with one percent Z-6040 during the dehydration processing stage showed no signs of resin separation. Thus, the use of Z-6040 throughout the dehydration stage is an effective method for improving the adhesion of Epon type epoxy resin to waxy biological tissue when processing samples for transmission electron microscopy. These results are important to electron microscopy experts, botanists, entomologists, and agricultural scientists in the government, at universities, and in private industry who are interested in obtaining high resolution images of plants and insects without tissue processing artifacts.

Technical Abstract: A commonly encountered obstacle when embedding waxy biological tissues for transmission electron microscopy, especially plant or insect tissue, is the tendency of the resin to detach from the tissue during ultramicrotomy. The inclusion of one percent Z-6040 organosilane (3-glycidoxypropyltrimethox-silane) during the dehydration process, functions as a coupling agent to enhance adhesion of the resin to the tissue. The use of Z-6040 for TEM was originally reported for embedding thin metal films and later adapted for embedding biological specimens in LR White acrylic resin. While LR White is a widely used resin for immunolabeling, it has several disadvantages and for morphological studies Epon-type epoxy resin can be a better alternative resin. Here we report on the use of Z-6040 as an additive during the tissue processing stage for embedding waxy biological tissue in Epon-type epoxy resin and examine the adhesion of the resin to eggs of the squash bug (Anasa tristis) and leaves of sweet orange (Citrus sinensis). Ultimately, samples embedded without the use of Z-6040 were observed to have small gaps along the perimeter of the cuticle where the resin had separated from the tissue. This separation can be especially problematic when studying fine details on the outermost cuticle of biological specimens. Samples which were primed with one percent Z-6040 during the dehydration stage showed no signs of resin separation. Thus, the use of Z-6040 throughout the dehydration stage is an effective method for improving the adhesion of Epon-type epoxy resin to waxy biological tissue, during the embedding process for transmission electron microscopy.