Secondary ion mass
spectrometry (SIMS) and confocal Raman microscopy
(CRM) are combined to analyze the chemical composition of cultured Pseudomonas aeruginosa biofilms, providing complementary
chemical information for multiple analytes within the sample. Precise
spatial correlation between SIMS and CRM images is achieved by applying
a chemical microdroplet array to the sample surface which is used
to navigate the sample, relocate regions of interest, and align image
data. CRM is then employed to nondestructively detect broad molecular
constituent classes—including proteins, carbohydrates, and,
for the first time, quinolone signaling molecules—in Pseudomonas-derived biofilms. Subsequent SIMS imaging at
the same location detects quinolone distributions in excellent agreement
with the CRM, discerns multiple quinolone species which differ slightly
in mass, resolves subtle differences in their distributions, and resolves
ambiguous compound assignments from CRM by determining specific molecular
identities via in situ tandem MS.