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Normalization of light element X‐ray intensities for surface topography effects in frozen‐hydrated biological bulk samples
Author(s) -
Marshall A. T.,
Condron R. J.
Publication year - 1985
Publication title -
journal of microscopy
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.569
H-Index - 111
eISSN - 1365-2818
pISSN - 0022-2720
DOI - 10.1111/j.1365-2818.1985.tb02665.x
Subject(s) - tilt (camera) , potassium , analytical chemistry (journal) , sodium , chemistry , absorption (acoustics) , detector , x ray , optics , materials science , chromatography , geometry , physics , mathematics , organic chemistry
SUMMARY X‐rays from light elements (Z=6–12) experience marked absorption in biological or organic matrices. Consequently surface tilt has a strong effect on X‐ray intensity. Using epoxy resin models with well‐defined surface tilt variations the effect of tilt on potassium, sodium and oxygen X‐ray intensities has been examined and methods for normalizing intensities for tilt effects have been investigated. With an energy dispersive X‐ray detector having a high (40°) take‐off angle, samples, having local tilt variations from 20° from the horizontal towards the detector to 20° away from the detector, can be analysed for potassium using peak intensities without loss of accuracy. Little loss in accuracy is experienced for sodium over the tilt range of 10° towards the detector to 10° away from the detector. Under these conditions peak to background (P/BG) ratios offer little advantage over simple intensity values. Sodium absorption in frozen‐hydrated biological samples is about 10% greater than in epoxy resin whilst potassium absorption is similar in both samples. It is assumed therefore that the foregoing conclusions also hold for frozen‐hydrated samples. X‐ray peak intensity to backscattered electron (P/BE) ratios provide a means of normalizing peak intensities from light elements for surface tilt effects. This is a particularly useful method for oxygen intensities because P/BG ratios cannot be obtained with any facility at the present time. Oxygen P/BE ratios provide a means of obtaining intracellular water concentrations in fractured frozen‐hydrated biological bulk samples, where surface tilt varies. Sodium intensities can also be normalized in this way.