Resolution:
## Figure 5.
Use of MIMS to study nitrogen-fixing bacteria.
(a-c) Secondary ion images from the molecular ions (a) ^{12}C^{14}N^{-}, (b) ^{12}C^{15}N^{-}, and (c) the HSI ^{12}C^{15}N^{-}/^{12}C^{14}N^{- }ratio of a sample containing both Teredinibacter turnerae (Tt; rod-like cells) and Enterococcus faecalis (Ef; bunches of rounded cells) cultured in a ^{15}N atmosphere for 120 h. Field: 46 × 46 μm (512 × 512 pixels); acquisition time 3 min.
The magenta color of the T. turnerae cells is an indication of their incorporation and fixation of ^{15}N (see Figure 3 for explanation). (d) The effect of scaling of the HSI ^{12}C^{15}N^{-}/^{12}C^{14}N^{- }ratio image (the numerator has been multiplied by 100) from T. turnerae cells exposed to a ^{15}N atmosphere for 32 h. Assigning the hue spectrum to the whole range of ratio values
allows easy identification of bacteria most highly enriched in ^{15}N (the turquoise cells in the top left panel). Compressing the hue scale (shown gradually
from top left to lower right) causes images of some of the cells to saturate at the
magenta level and allows us to easily recognize a succession of cells also enriched
in ^{15}N, although at a lower level. The isotope values start with 0–7 (top left; a value
of 7 is 19-fold higher than the natural ratio) and go to 0–0.5 (bottom right; a value
of 0.5 is 1.43 times the natural ratio). The field of view is 13 × 13 μm (256 × 256
pixels); acquisition time 20 min. (e,f) HSI image of the ^{12}C^{15}N^{-}/^{12}C^{14}N^{- }ratio (the numerator has been multiplied by 100) of a T. turnerae cell exposed to a ^{15}N atmosphere for 96 h. Field: (e) 8 × 8 μm; (f) 6 × 6 μm. Acquisition time: (e) 10
min; (f) 40 min. (g,h) HSI image of (g) the ^{12}C^{15}N^{-}/^{12}C^{14}N^{- }ratio (the numerator has been multiplied by 100) and (h) the ^{13}C^{-}/^{12}C^{- }ratio of T. turnerae in shipworm gill bacteriocytes incubated in the presence of a ^{15}N atmosphere for 4 h. Field: 10 μm × 10 μm (256 × 256 pixels); acquisition time 60
min. (i,j) HSI image (i) of the ^{12}C^{15}N^{-}/^{12}C^{14}N^{- }ratio (the numerator has been multiplied by 100) and (j) at ^{12}C^{15}N^{- }of T. turnerae exposed for 96 h in a ^{15}N atmosphere. Arrows indicate the flagella of the bacteria. Field: 60 × 60 μm (256
× 256 pixels); acquisition time 20 min. (k) Line scan across the flagellum observed in (i,j) showing ^{12}C^{15}N^{- }secondary-ion counts as a function of pixel address across the flagellum. One pixel
is equivalent to 234 nm. Inset: arrow points to the flagellum; the red box indicates
the area of the bacterium that was used to evaluate the mean ^{12}C^{15}N^{- }counts.
Lechene |