Histochemical methods for the detection of apoptosis in the nervous system

Neuroscientists often need to detect neuron death at the light microscope level in tissue sections derived from animal models of neurological disease. In many instances there is a need to detect apoptosis, the most common morphology of programmed cell death. This unit provides two protocols for the detection of apoptosis by immunostaining for either activated forms of caspases or their cleavage products. When used in conjunction with nuclear dyes, these protocols permit visualization not only of caspase activation, but also the nuclear chromatin clumps characteristic of apoptosis. The first protocol utilizes peroxidase-mediated chromogen deposition to visualize antibodies by brightfield microscopy. The second protocol utilizes fluorophores to visualize antibodies by epifluorescence. Double immunofluorescence labeling can be performed to identify the phenotype of cells in which caspases are activated. Not all cell death is apoptotic. Therefore, a third protocol is presented for suppressed silver staining, a useful method to screen for all morphologic forms of cell death.