Table 2 |.

Troubleshooting table

Step	Problem	Possible reason	Solution
2–4	15N-labeling not detected	Inadequate label dose or delivery	Use controls to assess for absorption (urine) and/or incorporation into alternative cell types (WBCs)
		Label is diluted away by ongoing proliferation during label free chase	Increase dose of 15N-thymidine labeling or decrease chase period
27–40	Indeterminate cell type in NanoSIMS images	Inadequate resolution or edge effect at margin of imaging field	Reanalyze with NanoSIMS at higher resolution and with centering of the histologic feature of interest in the imaging field
	Cannot resolve mass peaks for the N isotopes	The MRP is not high enough	Iteratively adjust Q, LF4, HEX and incrementally use smaller entrance and aperture slits
			Insert the smallest exit slit (ExS 3) for the detector monitoring 15N
36	Long pre-sputtering time	Primary ion beam intensity is too low	Adjust the voltage (higher or lower) slightly on the L1 primary lens, in coordination with deflection plates C1x and C1y
	Secondary ion intensity unstable	Sample is not flat and/or smooth	Inspect sample height with a microscope and readjust sample in NanoSIMS holder Apply a thicker gold coat
55	Cardiomyocytes in oblique axis for binucleation quantification		Review additional serial sections in the z-axis (sections are on glass slides) for a second nucleus
66	Over- or undersaturation of photomicrographs		Decrease concentration of Hoechst staining or incubation time
			Adjust the strength of the excitation light, the sensitivity of the emission light detector, and the exposure time