4D defocused particle tracking (DPT) technique

As we enter the era of quantitative biology, there is an immediate need for a new set of quantitative tools to probe the spatial and dynamical information of biological entities (such as bacterial cells, proteins)  at micro and nano scales. We have developed imaging techniques that are able to track micron meter scale particles  in all four  dimensions (x,y,z,t) using a single CCD video camera.


The idea behind this imaging technology is straightforward. When a point like object is in focus, one gets a focused point in the imaging plane; when a point like object moves away from the focal plane, one gets a fuzzy dot in the imaging plane. The graph below shows images of a 1 micron fluorescent bead at different vertical positions using an epi-fluorescent microscope. As seen the defocused image contains all the information necessary to infer the (x,y,z) location of the fluorescent beads.

Fig. 1 Defocoused images of a 1um diameter fluorescent bead at different z locations.


Here are a few successful examples using the 4D DPT technology


Tracking swimming E.coli in four dimensions.

Revealing Lagrangian flow fields in a micro mixer.


If you are interested in using our automatic tracker, please contact Mingming Wu at mw272@cornell.edu. You can also

download the tracking software at 4D-Tracker.  This is a series of program that (1) translates images with defocused particle images into (x,y,z,t) coordinates; (2) forming trajectories by connecting particles of nearest neighbors.



  1. Mingming Wu, John Roberts, and Mark Buckley, Three-dimensional fluorescent  particle tracking at micron-scale  using a single camera,  Experiments in Fluids, 38, 461-465(2005).
  2.  Mingming Wu and Morteza Gharib,  Experimental studies on the shape and path of small air bubbles rising in clean water, Physics of Fluids, 14, L49(2002)

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