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. 2023 Jul 31;13:12383. doi: 10.1038/s41598-023-38699-1

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© The Author(s) 2023

Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons licence, and indicate if changes were made. The images or other third party material in this article are included in the article's Creative Commons licence, unless indicated otherwise in a credit line to the material. If material is not included in the article's Creative Commons licence and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this licence, visit http://creativecommons.org/licenses/by/4.0/.

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Mass spectrometry imaging of DISC-3D samples (10 μm slices) of MDA-MB-231 spheroids. Average intensity from mass spectrometry signals averaged over spheroid sections at various temperatures across (a) the full 200–1000 m/z range, (b) the 300–500 m/z range, and (c) the 700–900 m/z range. Statistical results shown here were conducted as pairwise samples. A full description of statistical results is presented in Fig. S6b. n = 8 in all cases except 150 °C, where n = 7. *p < 0.05, ^†p > 0.05. (d,e) Representative normalized mass spectra collected from invasive spheroids following the DISC-3D protocol at (d) 24 °C and (e) 350 °C demonstrating the increase in signal intensity in the ≈ 650–900 m/z regime. Number at right on each panel indicates maximum intensity peak at that temperature. (f) Representative mass spectrometry images prepared following the DISC-3D protocol outlined in Fig. S1d, showing consecutive sections of single spheroids for several signal ions. In the first serial section of each signal ion, the spheroid core is marked by a white dotted line and the invasive front is denoted by a red dotted line. Color scale from minimum to maximum intensity is shown for signal ions at right. Scale bar = 500 μm.