The diversity of insect neuropeptides coupled with the limitations from the small size of the insects themselves combine to make positive identification through peptide sequencing a highly challenging task. The advent of the “soft-ionisation” techniques of MALDI-TOF and electrospray (ESI)-Q-TOF mass spectrometry, coupled with the additional information from insect genome projects have revolutionised the characterisation of insect neuropeptides, such that sequences can now be obtained from just a few cells, where before thousands of insects had to be laboriously dissected, extracted and purified. Some of the procedures that are now used to identify these peptides are described here. Once the neuropeptides have been identified, it then becomes possible to use this knowledge to define physiological functionality.