The Hidden Power of Coffee – An Exceptional Replacement for Uranyl Acetate for Staining Biological Specimens in Transmission Electron Microscopy

The use of uranyl acetate, one of the most conventional contrast agents used for decades inelectron microscopic examinations of biological specimens, is strictly regulated by law and in somecases banned from laboratories. In recent years, non-toxic or non-radioactive alternatives withsimilar staining results have been developed. These alternatives are not equally effective for allbiological structures. Examples are Uranyl Acetate Replacement (UAR), Uranyless (UL) andOolong tea extract (OTE) [1, 2]. In this work, two new alternatives, being coffee solution as well asone of its components, chlorogenic acid (CGA), have been studied [3].Coffee contains a wide range of ingredients that are said to have health-promoting traits. A newoutstanding but unknown property is that coffee can also be used to increase contrast of biologicalsamples in electron microscopy. Figure 1 shows mitochondria in zebrafish, A) stained with UA, B)stained with coffee. Subjectively, both images show quite good contrast. This work illustrates howa subjective impression of either good or poor contrast can be converted into an objective andthus comparable numerical value using the Michelson contrast and a MATLAB algorithm (see Fig.2), which makes a direct comparison of the different contrast agents possible. On comparing thesecontrast values for epithelial cells, cell membranes, muscle fibers tubules and mitochondria inzebrafish it turned out that coffee is even more suitable as a contrast agent than UA. Alsochlorogenic acid, for its part, produces good image quality with high contrast values and issuperior to UA concerning mitochondria, cell membranes, muscle fibers and epithelial cells. Thismethod now allows the direct numerical comparison between differently stained specimens. Forthis research, primarily the excellently mapped zebrafish (Danio rerio) was used, but subsequentlymouse kidney was also studied in detail.