these types of imaging based approaches, it is possible to combine these methods with other single cell gene expression quantification methods such as RNAseq. Tracking Tracking is another traditional image processing problem which appears in bioimage informatics. The problem is to relate objects that appear in subsequent frames of a film. As with segmentation, the problem can be posed in both two- and three-dimensional forms. In the case of fluorescent imaging, tracking must often be performed on very low contrast images. As obtaining high contrast is done by shining more light which damages the sample and destroys the dye, illumination is kept at a minimum. It is often useful to think of a photon budget: the number of photons that can be used for imaging before the damage to the sample is so great that data can no longer be trusted. Therefore, if high contrast images are to be obtained, then only a few frames can be used; while for long movies, each frame will be of very low contrast. Registration When image data samples of different natures, such as those corresponding to different labeling methods, different individuals, samples at different time points, etc. are considered, images often need to be registered for better comparison. One example is as time-course data is collected, images in subsequent frames must often be registered so that minor shifts in the camera position can be corrected for. Another example is that when many images of a model animal (e.g. C. elegans or Drosophila brain or a mouse brain) are collected, there is often a substantial need to register these images to compare their patterns (e.g. those correspond to the same or different neuron population, those share or differ in the gene expression, etc.). Medical image registration software packages were early attempts to be used for the microscopic image registration applications. However, due to the often much larger image file size and a much bigger number of specimens in the experiments, in many cases it is needed to develop new 3D image registration software. The BrainAligner is a software that has been used to automate the 3D deformable and nonlinear registration process using a reliable-landmark-matching strategy. It has been primarily used to generate more than 50,000 3D standardized fruitfly brain images at Janelia Farm of HHMI, with other applications including dragonfly and mice. Important Venues A consortium of scientists from universities and research institutes have