PROJECT SUMMARY Three-dimensional (3D) microscopy offers many promises for biological investigations and medical applications. However, it is currently limited to well-resourced laboratories in settings with established infrastructure. Many 3D microscopy techniques (e.g., confocal) rely on focusing light at an array of small locations within the sample, which requires expensive and specialized equipment. Optical Projection Tomography (OPT) is a 3D imaging technique that utilizes traditional microscopy equipment; instead of focusing the light at specific locations, OPT images a sample from many angles to reconstruct a 3D volume. It is a very effective method for 3D imaging of small translucent objects (e.g., mouse fetuses, parasites, and large bacteria). While it is possible for OPT to be a lower-cost method of 3D microscopy, existing systems remain expensive and large. We propose to take advantage of the ubiquitous and high-quality computing and imaging hardware available in smartphones to make an OPT device that is inexpensive and extremely portable. We will create a smartphone extension that robustly images a rotating sample and uses the phone’s computational hardware to reconstruct the 3D volume. Two imaging modalities will be pursued: visible-band attenuation microscopy (e.g., brightfield) and luminescent microscopy (e.g., bioluminescent). The components of the smartphone extension are either 3D printed, laser cut acrylic, or readily commercially available. Thus, the cost of manufacturing the device is extremely small and the total weight of the device is very low; the total cost of all components will be less than $50. The components of the extension are easily assembled on site, permitting the device to be transported with in a small package. Due to its low cost and size, the OPT microscope can also serve as a useful tool for educational purposes (e.g., as part of an undergraduate laboratory course involving optics) and for generating real data for tomographic algorithm development. To validate the device, we will build two phantoms with three-dimensional features that allow us to evaluate the Modulation Transfer Function: one for attenuation microscopy and another for luminescent microscopy. Aim 1: Build a visible-band tomographic microscope extension to a smartphone. Aim 1A: Implement 3D cone-beam reconstruction from visible-band sinogram data for samples approximately 10 mm in size with approximately 10 µm resolution. Aim 1B: Improve the image resolution with a multi-lens optical system. Aim 2: Implement bioluminescent tomographic microscopy by appropriately modifying the tomographic reconstruction algorithm. Aim 3: Build two shelf-stable 3D phantoms with features of sizes varying from 1 µm to 200 µm.