Fate Mapping and Transcript Profiling of Germinal Center Cells by Two-Photon Photoconversion.

The germinal center (GC) reaction is the key process for the generation of high affinity antibodies to foreign antigen. Standard experimental techniques such as fluorescence-activated cell sorting and histology have provided numerous insights into the composition and function of the GC. However, these approaches are limited to a "snapshot" in time and are unable to fully capture the dynamic nature of the GC. Intravital two-photon microscopy overcomes these disadvantages and has led to several major advances in the field but is restricted by practical and technical limits that prevent long-range mapping and molecular studies. Here we describe procedures for optical marking or "tagging" of cells in precise microanatomical compartments by two-photon photoconversion that can be used for long-term fate mapping and transcript profiling of GC T and B cells.