Thioflavin T Stain: An Easier and More Sensitive Method for Amyloid Detection

In this chapter, the application of thioflavin stains to the diagnosis of amyloid is presented. Although Congo red stain is considered to be the generally accepted standard for the diagnosis of amyloid, it requires expertise and the use of polarized light microscopy; moreover, the diagnostic “apple green birefringence” of the dye may be difficult to visualize and, therefore, show low sensitivity. Congo red is a direct dye with different affinities for fibrillar and nonfibrillar materials. Therefore, the protocol involves staining followed by washing, and this may lead to significant levels of background stain and lower reproducibility. In contrast, the fluorochrome dyes, thioflavin S and T, exhibit a dramatic increase in fluorescent brightness upon binding to amyloid. The binding of thioflavin dyes is linked to the presence of cross-β structure in the amyloid fibrils, and this stain is very sensitive. However, thioflavin stains are less specific for amyloid than is the Congo red stain. Therefore, it is preferable to confirm the results obtained with thioflavin by either electron microscopy or the Congo red stain itself. Nevertheless, given the ease of staining, the predictability of stain outcome, and the ease of interpretation, adding thioflavin to the staining repertoire may be a welcome solution to problems that could arise from the use of Congo red stain alone in the clinical diagnosis of amyloid.