In 2007, probe electrospray ionization/mass spectrometry (PESI/MS) was developed. In this technique, the needle is moved down along a vertical axis and the tip of the needle touched to the sample. After capturing the sample at the needle tip, the needle is then moved up and a high voltage is applied to the needle at the highest position to generate electrospray. Due to the discontinuous sampling followed by the generation of spontaneous electrospray, sequential and exhaustive electrospray takes place depending on the surface activity of the analytes. As modified versions of PESI, dipping PESI (dPESI), sheath-flow PESI (sfPESI) and adjustable sfPESI (ad-sfPESI) have been developed. These methods are complementary to each other and they can be applicable to surface and bulk analysis of various biological samples. In this article, the characteristics of these methods and their applications to real samples will be reviewed.