In order to investigate metabolic properties of single cells of freshwater algae (Haematococcus pluvialis), we implement matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS) in combination with microspectroscopic mapping. Straightforward coupling of these two detection platforms was possible thanks to the self-aliquoting properties of micro-arrays for mass spectrometry (MAMS). Following Raman and fluorescence imaging, the isolated cells were covered with a MALDI matrix for targeted metabolic analysis by MALDI-MS. The three consecutive measurements carried out on the same cells yielded complementary information. Using this method, we were able to study the encystment of H. pluvialis-by monitoring the adenosine triphosphate (ATP) to adenosine diphosphate (ADP) ratio during the build-up of astaxanthin in the cells as well as the release of β-carotene, the precursor of astaxanthin, into the cytosol. © 2013 The Royal Society of Chemistry.