Synthetic Vesicle Size Analysis

Background

I have a few hundred of these cryo-em pictures of synthetic vesicles.
They are all roughly spherical and I just need to measure their diameters. The large structure that dissects the image into three parts is just the TEM grid and should be excluded from the analysis.

1.tif (1.0 MB)

Challenges

I’m having issues detecting the edges of the vesicles since they are only a slightly darker grey and are only a few pixels wide.

I’m using Fiji but am new at the automation. Plugins like particle analyzer can’t seem to differentiate the vesicles from the background no matter how I set the threshold. I also tried diameterj, but it also can’t seem to detect the edges of the vesicles apart from background.

I also looked into trying to use cell profiler but don’t know where to start as far as the image processing side of things goes.

Thank you for any help!

Hi,

if your vesicles are always expected to be circular, you might get it to work using a combination of the Ridge detection plugin and the Hough Circle transform in Fiji.

Although especially the circle detection takes quite some time to run per image, I was surprised by the results given the low contrast of the vesicles.

run("Subtract Background...", "rolling=5 light sliding");

// ridge detection
run("Ridge Detection", "line_width=5 "+ 
"high_contrast=230 low_contrast=87 " +
"darkline correct_position extend_line "+
"make_binary method_for_overlap_resolution=NONE sigma=2 "+
"lower_threshold=0.34 upper_threshold=3.23 minimum_line_length=0 maximum=0");

// Hough Circle Transform 
run("Hough Circle Transform",
	"minRadius=15, maxRadius=200, "+ // expected min and max radius of circles in px
	"inc=1, minCircles=1, maxCircles=100, "+ // expected max number of circles per image
	"threshold=0.3, "+ // threshold between 0 and 1 to adjust sensitivity of detection
	"resolution=1000, ratio=1.0, bandwidth=10, local_radius=10,  "+
	"reduce show_mask results_table");

If you play around with the settings in the GUI-versions of the respective plugins some more, you might get an even better detection. Just make sure you turn on the Macro recorder.

Good luck, Joachim

3 Likes

Wow… I’ve been playing around with it for two nights and wasn’t able to get that far. I’ll experiment with these plugins and post back if I run into any major issues.

Thank you so much!