How Do I Analyze My Experiment?

Image of yeast cells magnified 20xIn a typical live cell experiment, many locations are taken in a specific well and many wells are included in the experiment. The first important thing to consider is how many images should I take in a given well, then how many replicates should I have to make sure I am seeing a given effect. That will be covered in a future write up on statistics. But in any case, a typical experiment will last many days with each location producing a series of images, typically every 10 to 15 minutes between images.

The first way to look at the results of an experiment is to just look at time lapse images using any image viewer. That will give you a sense of the experiment – did the cells grow, do they look healthy, do you see any general trend. The next step is to produce an AVI of some interesting events in the experiment. For this process we recommend an open source package, ImageJ. ImageJ will run on most computers, it’s been around since 1997 and has many simple and advanced image processing operations.

To load a sequence of images in ImageJ, just select File>Import>Image Sequence…, then locate the starting image for the sequence. The program will load all the images. You can view the image sequence and can also save an AVI of the sequence, select Save As>AVI. You can now send this AVI to anyone or store the sequence as a single file. ImageJ will also load an AVI and separate out the images. You can use this to rename the images. There are many other operations available in ImageJ as well as an open source package called Fiji. This is just a version of ImageJ with many of the options called plugins already installed.

Another convenient way to look at an image sequence is to create a montage of the sequence. Open Image>Stacks>Make Montage. This operation will produce a single image of the complete experiment, a good way to look for any problems during the experiment or to see any general effects like cell growth for example. This image can also be saved for future reference. A typical AVI of 100 images might be 6MB where a montage is only 0.5MB. ImageJ will also allow you to produce macros for repeated operations and many image processing operations. Refer to resources on our web site, to get some simple instructions on how to count and track cells using ImageJ or Fiji.

Here is an example of a Montage created in ImageJ:

( Look for lost images, lighting issues and most important cell growth ! )


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