X-Ray Microanalysis Techniques

X-ray production is often the inevitable consequence of the electron irradiation of atoms, whether the atoms reside in biological or materials specimens that are either thin or infinitely thick. As long as the incident electrons are invested with sufficient kinetic energy to ionize individual atoms, X-rays will emanate from them. The technique of biological X-ray microanalysis, henceforth generically referred to as EPXMA (electron probe X-ray microanalysis), has been evolving for over 25 years, and its principles and practicalities have been described in detail on a number of occasions over that period (see [1 -5 ] for primary sources). It is a technique that detects and measures elements within structurally defined “compartments”; it cannot distinguish “bound” from “free” elemental pools; it cannot differentiate between isotopic or redox states. The spatial resolution of EPXMA is unsurpassed by any other analytical technique that combines simultaneous compositional and morphological observation. The sensitivity of EPXMA seems impressive (10-¹⁸ to 10-¹⁹ g of an element like Ca under favorable conditions), until this is converted into a concentration value for the local analyzed compartment (about 2 mmoles/kg weight, or 200 μg/g , for Ca), which is several orders of magnitude poorer than, for example, “bulk”s analytical techniques such as atomic absorption spectrophotometry and induction coupled plasma analysis. It is worth bearing these basic facts in mind before embarking on a tortuous EPXMA study, whilst also acknowledging that trace elements may not necessarily be homogeneously distributed through a biological system: they may be localized within a certain cohort of specialized cells and/or sequestered within discrete subcellular compartments. The limiting factor for EPXMA detection is local not bulk concentration.