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![[Figure 1]](wuz5001fig1mag.jpg)
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Figure 1
Sample preparation workflows and imaging principle for phase-contrast imaging. (a) Schematic of X-ray propagation through resin-embedded, heavy-metal-stained neuronal tissue. The difference in refractive index between tissue and surrounding resin causes modest distortion of the X-ray wavefront. z1 and z2 are the distances from the X-ray source or focal spot to the sample, and from the sample to the detector, respectively. (b) Equivalent schematic for critical point dried (CPD) tissue imaged in air or vacuum. The absence of embedding resin introduces a stronger refractive index discontinuity at the tissue–air interface, leading to enhanced wavefront distortion and increased phase contrast. (c, d) Standard preparation pipeline for Epon embedding (c) and CPD preparation (d), both following fixation, staining, and dehydration. (e, f) Representative single-projection images of femtosecond-laser-milled cylindrical pillars imaged at 17 keV in air: Epon-embedded (e) and CPD-prepared (f). (g)–(i) CPD workflow: (g) ethanol-to-CO2 exchange gradient during CPD processing, (h) phase diagram of CO2 indicating the critical point transition, and (i) illustration of the final CPD step where liquid CO2 is brought to the critical point and then slowly vented as gas while maintaining temperature, avoiding surface-tension forces associated with a liquid–gas interface and preserving ultrastructure (Williams & Clifford, 2000  ). |
![[Figure 1]](wuz5001fig1.jpg)
 | JOURNAL OF SYNCHROTRON RADIATION |
ISSN: 1600-5775
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