Optical scatter imaging as an apoptosis assay for cells undergoing ALA-mediated photodynamic therapy in vitro

Vincent M. Rossi, Paige J. Baugher, Steven L. Jacques

Research output: Chapter in Book/Report/Conference proceedingConference contribution


Aminolevulinic acid (ALA) is converted to protoporphyrin-IX (PpIX) within mitochondria, causing the assumption that ALA-mediated photodynamic therapy (PDT) results in mitochondrial damage and therefore an apoptotic response. Mitochondria within apoptosing cells swell, forming pores in their outer mitochondrial membranes which release cytochrome-c, triggering apoptosis. Optical scatter imaging (OSI) makes use of scattered fields in order to indicate the morphology of subcellular components, and is used here in order to measure changes in mitochondrial size as a response to ALA-mediated PDT. Two images of the same field of view are spatially filtered in the Fourier plane of a 4-F system. Both spatial filters block directly transmitted light, while accepting different angles of scattered light through an adjustable iris. The optical scatter image ratio (OSIR) of the local intensities of these two spatially filtered images is indicative of scattering particle size. Mie theory is used to calculate the predicted OSIR as a function of scattering particle size. In this fashion, the OSI system is calibrated using polystyrene microspheres of know sizes. Comparison of the measured OSIR from cellular images to theoretical values predicted for mitochondria then serves as an indication as to whether cells are apoptosing. Cells are treated at varying concentrations of ALA and varying exposures of 635 nm light and imaged at varying time points in order to develop a broader understanding of an apoptotic response of cells undergoing ALA mediated PDT.

Original languageEnglish (US)
Title of host publicationBiomedical Applications of Light Scattering VII
StatePublished - 2013
Externally publishedYes
EventBiomedical Applications of Light Scattering VII - San Francisco, CA, United States
Duration: Feb 2 2013Feb 4 2013

Publication series

NameProgress in Biomedical Optics and Imaging - Proceedings of SPIE
ISSN (Print)1605-7422


OtherBiomedical Applications of Light Scattering VII
Country/TerritoryUnited States
CitySan Francisco, CA


  • MG-63 cells
  • Optical Scatter Imaging
  • PpIX
  • aminolevulinic acid
  • mitochondria morphology
  • osteosarcoma
  • photodynamic therapy

ASJC Scopus subject areas

  • Electronic, Optical and Magnetic Materials
  • Biomaterials
  • Atomic and Molecular Physics, and Optics
  • Radiology Nuclear Medicine and imaging


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