Protocol for reconstructing 2D and 3D peptide atlases of the Ciona cerebral ganglion at single-cell resolution

The distribution of biomolecules is essential information for understanding biological systems. Here, we present a protocol for 2D and 3D reconstruction of tissue images at the single-cell level by integrating microscopy and mass spectrometry imaging (MSI) data through linear interpolation. We describe steps for sample preparation, MSI, and construction of 2D and 3D single-cell peptide atlases of the cerebral ganglion of Ciona robusta. This approach is also applicable to other animals in which target cells are labeled with endogenous fluorescent proteins. For complete details on the use and execution of this protocol, please refer to Osugi et al.¹.