A representative H&E-stained section and MS images of peptide and protein standard mass spectra from serial sections of a single adult, female ZF embedded in 2% CMC + 5% gelatin and sprayed with three different matrices in 50% ACN, 0.1% TFA. (A) H&E-stained serial section; (B) 5 mg/ml SPA, mass standard apomyoglobin+2, average mass 8476.78 Da; (C) 5 mg/ml CHCA, mass standard angiotensin I, monoisotopic mass 1296.6853; (D) 5 mg/ml CHCA + 6 mM AmP, mass standard angiotensin I, monoisotopic mass 1296.6853. The standard mass for which the droplet is highlighted in the MSI is labeled on the mass spectrum. Arrows in the spectra denote appearance of expected mass-standard peak; a circle in the MSI denotes mass-standard droplet placement; overall average mass spectrum for the circled area and total ion current is shown for each mass spectrum.