Neurodegenerative diseases such as Alzheimer’s or Parkinson’s are marked by progressive loss of affected neurons. Even with novel disease-modifying therapies, this loss cannot be reversed. In situ astrocyte-to-neuron (AtN) transdifferentiation may provide an opportunity to convert resident astrocytes into new neurons to revert the loss of neurons incurred. Currently, most studies investigating AtN transdifferentiation in vitro rely on the use of primary mouse astrocyte cultures which require sacrificing animals and come with uncertainty regarding species differences. Conversely, human induced pluripotent stem cell (hiPSC)-derived astrocytes offer the advantage of working in a human cell culture system which improves translatability and provides the opportunity to generate large, cryopreservable batches of cells to identify and study conversion factors. This protocol details a workflow for assessing the suitability of potential conversion factors for transdifferentiating hiPSC-derived astrocytes into neurons.