Fig. 1
SRS microscopic imaging of biosynthetic incorporation of deuterium from D2O into macromolecules in living cells and animals. a D2O-derived deuterium can form O–D, S–D, and N–D bonds through reversible non-enzymatic H/D exchange and be incorporated into C–D bonds of metabolic precursors for the synthesis of macromolecules through irreversible enzymatic incorporation. Note that, under the condition of sparse labeling, the position of deuterium labeling on lipids is random, and this illustration only represents one possible labeling pattern. b Various X–D bonds produced Raman peaks at distinct positions. C–D, C–H, and O–D spectra were collected from Hela cells grown in 70% D2O medium, S–D spectrum from saturated cysteine dissolved in D2O, and N–D spectrum adopted65. SRS microscopic images were collected for C–D, O–D, and C–H signal of HeLa cells at 5 min and 24 h after adding D2O-containing medium. c Signal-to-noise ratio (S/N; noise = 1 μV) for SRS microscopic signal of sebaceous glands at 2135 cm−1 from mice that drank 4% or 25% D2O for 8 days. Percentages of D2O enrichment in body water are shown in parentheses. Detection limit at S/N = 10 or 2 was calculated based on the linear relationship between average S/N and D2O enrichment level. d, e Frames from live SRS microscopic imaging recordings of the sebaceous glands under the ear skin of intact mice that drank 25% D2O for 9 days (Supplementary Movie 1–3) and living fourth stage C. elegans larva that grew on 20% D2O-containing NGM plates for 4 h (Supplementary Movie 4–6). The blood flow (orange arrows) in mouse visualized with two-photon absorption contrast and the movement of the C. elegans body (white arrows) indicated that the animals under imaging were alive. Scale bar = 20 μm. C–D signal was shown in cyan hot LUT, and color scale bars display the scale for the lookup table values
