Fig. 1: MEZ-XRF principle and element detection limits.
From: Multielement Z-tag imaging by X-ray fluorescence microscopy for next-generation multiplex imaging
a, MEZ-XRF involves (1) staining of biological samples with Z-tagged affinity reagents, (2) raster scanning of a focused X-ray beam over the stained sample and collecting emission spectra for each pixel, (3) deconvoluting spectra into multichannel images and (4) analyzing images. Resolution is determined by the focus and step size of the raster X-ray beam. Repeat imaging at different resolutions enables multiscale MEZ-XRF. b, The highest yield L- and K-line emissions for the 20 elements used as Z-tags and stable isotopes used for isotope-tagged reagents. All elements were included in the multielement gelatin standard except those in italics. Elements indicated in bold have a single isotope enabling direct comparison between XRF and IMC signal. c, Averaged 1-s emission spectra (n = 20), triggered by a 69-keV X-ray beam raster scanned over a gelatin standard containing 200 ppm of each Z-tag element (except Rh and I). XRF emissions were recorded with an SDD (black) or GeCMOS (blue) detector. The major Kα1 emission lines of Z-tag elements are labeled. d, XRF detection limits determined from an eight-point serial dilution series for each element in the multielement gelatin standard as measured by deconvoluted Kα1 emissions (#Lα1 for Ir) with the indicated detectors and raster rates. e, IMC detection limits determined from eight-point serial dilution gelatin standards with raster scanning of 500 nm2 pixels at 200 Hz. IMC signals from an identifiable isotope of a single element are labeled. Unlabeled lanthanide isotopes are mixtures of isotopes (b) that are isotope abundance-adjusted according to their presence in the single-element standard solutions. IMC isotope abundances per element are shown in Extended Data Fig. 3.
