Imaging unstained proteoglycan aggregates by soft x-ray contact microscopy

Soft X-ray contact microscopy is a relatively new form of ultrastructural imaging, having better than 6 nm resolution and being uniquely well suited for the examination of fragile, unstained biological specimens. The biological specimen placed on a layer of photoresist and exposed to soft X-rays (1-10 nm lambda) of a specific wavelength or broad band. After X-ray exposure, the specimen is removed from the photoresist and the latter chemically developed. When the developed replica is examined by high resolution scanning electron microscopy, the fine structure of the original biological specimen is faithfully reproduced. Since the soft X-ray replica is initially formed due to the differential absorption of the incident X-rays by the biological specimen, the resultant contact replica also reveals information about the elemental composition of the sample. This paper presents our application of this new technique for the study of the proteoglycans, the complex polyanionic macromolecules comprising the gel phase in the matrix of mammalian cartilage.