As might be expected, there are also differences in mass spectra obtained by direct ionization using MAI, MALDI, and ESI. The differences are best seen when comparing complex mixtures. One example is the comparison of ESI and MAI using the automated system which is capable of analyzing 384-well microtiter plates or surfaces, for example glass plates, using ESI, MAI, SAI, or vSAI, just as with the manual system. Three strains of E. coli bacteria were grown, lysed, extracted in ethanol, and five replicates of each extract were acquired by MS using MAI, with the 3-NBN matrix in 3:1 acetonitrile–water, as well as with ESI directly from the ethanol extract. The results for the m/z range 400–800 of MV1184 E. coli in ethanol are shown in Figure 2. Some of the differences observed are ionization by Na + adduction in ESI versus protonation in MAI, for example m/z 726 and 704, while other ions, for example m/z 749 and 765 in ESI and ions around m/z 637 in MAI, are not observed in both mass spectra. Because protonated ions fragment more readily, the MAI mass spectrum shows mass differences of m/z 141 ( m/z 690 to 549 and m/z 704 to 563), likely representing loss of the polar head group from a glycerophospholipid. Thus, some of the contents of this extract as viewed by these two ionization methods would seem to be different.