Heterogeneity in cell populations is a phenomenon encountered in virtually all biological systems; in whole tissues and cell cultures, and from bacterial to mammalian cells. Moreover, assessment and correct interpretation of these cell-to-cell differences is increasingly being recognized as an important factor in biology.1 To study cell heterogeneity in phenotype, genotype and their chemical products, new analytical methods that allow the examination of individual cells are constantly being developed. These methods are often characterized by their exquisite spatial and temporal resolution, as well as extreme sensitivity. They often find application in biochemical research, clinical diagnostics, drug development, regenerative medicine and tissue engineering. This review aims to provide an overview of the technological advances made in this still rapidly expanding field during the last two years. We will focus on new developments or variations of existing techniques in three major areas of quantitative chemical analysis: electrochemical analysis, optical analysis and (super resolution) microscopy, and mass spectrometry and mass spectrometry imaging. Some relevant examples of qualitative analysis will be discussed as well. Techniques are increasingly applied in a hybrid fashion in an effort to answer evermore complex biological questions as the importance of differences between individual cells is better understood and recognized. Given the extent to which many developments rely on microfluidics, a dedicated section on this is included. Some of the many recent examples of single-cell "omics" performed in microfluidic devices are also discussed. In an effort to write a comprehensive yet not utterly exhaustive overview, we have decided to leave out other areas that have not seen as many new developments. We have also decided to omit techniques in the major areas that have become established and standard approaches, unless there have been significant new developments. We sincerely hope we have paid attention to all the work done in this field and apologize to anyone whose work we omitted.