I am taking time lapse videos of cells using a confocal microscope, and we want to quantify the average cell length of cells at specific times and generate a histogram of cell lengths. So I need to threshold, and then separate growing rod shaped E. coli cells, then quantify the cell length of all the cells in the image. I have been using FIJI, and I know how to quantify the length of a cell but only one at a time and this would be very slow for hundreds of cells over multiple experiments. Does anyone know of a pipeline, macro, or plugin to accomplish this easily and reliably?
Thank you
Hi Chris Hamm ,
You should look into manuals how to use the particle analyser to count cells/objects. Than you will find this site interesting:
https://imagej.nih.gov/ij/docs/menus/analyze.html
There you will find the different measurements explained that you can apply in the particle analyser. I think the best for your problem would be the Feret's Diameter if your cells are fusing into clumbs of multiple cells you might want to try a water shed filter first.
Best wishes
Soenke
Dear Chris Hamm as you say you would have to use thresholding to get a good mask separating your cells and corresponding well to their outline. This is the most tricky part. You can do that for all the images together if you generate a stack with all of them (using stack histogramm in the threshold option) but as Sönke mentions u might need to refine using watershed or other corrections. After u have a good segmentation, u can define all parameters u want to measure (analyse/set measurements; i believe u need min.feret and multiply it with aspect ration (in shape descriptors?) to get the length, but not 100% sure) and then you can get the data for all the cells (analyse/analyse particles) u need to tell it to keep the label in set measurements "show label" so that u know from which image of the stack it came, and then if u analyse particles it asks you to include all images and u say yes -> thus you should get all the data you need. Or u can automate it for a batch of images in a folder. Good luck!
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