Usually I did it with imaje J. To get statistic around 100 particles in one SEM image take around 5-10 minutes.

Briefly:

open image with Image J.

1) set up the scale. (draw line along your scale and in menu Analyze>set scale. put you scale size.

2) back to image, use line tool to along the particle diameter. press the M button.

3) repeat it (2) with all particles and as result you get a table with all sizes.

4) you can get the mean and SD by Analyze>Distribution

ImageJ is quite a good freeware for this type of analysis.

If you have labview you can also use its graphics add-on to analyze pictures.

just follow this link.

- Download ImageJ: http://rsbweb.nih.gov/ij/download.html

- Tutorial and documentation: http://rsbweb.nih.gov/ij/docs/index.html

- Plugins: http://rsbweb.nih.gov/ij/plugins/index.html

I used Gwyddion. Its more for AFM but it can also measure the size of particles on ur SEM image

I agree with Mr. Guillame ImageJ is so ggod for ur inquiry Mr. Behzad

Usually I did it with imaje J. To get statistic around 100 particles in one SEM image take around 5-10 minutes.

Briefly:

open image with Image J.

1) set up the scale. (draw line along your scale and in menu Analyze>set scale. put you scale size.

2) back to image, use line tool to along the particle diameter. press the M button.

3) repeat it (2) with all particles and as result you get a table with all sizes.

4) you can get the mean and SD by Analyze>Distribution

Thank you all.

This is manual measuring. Isn’t there any software for automatically measuring? For example by selecting a specific area, it gives us mean particle size.

Mountains map from Digital Surf and SPIP from Image Met can do this, But unfortunately this is not free !!

Thank you Sébastien.

What about agglomerated or overlapped particles? Can we measuring them by Gwyddion or ImageJ?

better do TEM to measure particle size exactly. In SEM you will get agglomeration of NP.

You can use image J for it. But I think SEM will not be good tool for nanoparticles in case if your particles are less than 50nm. May be you try TEM for better results

Thank you Arun and Naeem.

Dear Naeem,

Would you tell me how can I measure agglomerated particles by ImageJ?

Are the plugins of ImageJ software required for particles measuring?

Automatic particle analysis is included in std ImageJ. In case of agglomerates, it might not work as well. You should try to improve your sample prep in order to get better images, rather than rely on some software magic though.

"off-topic" ... to comment on the SEM data recording:

Depending on the size of the nanoparticles imaging the particles in transmission even in a scanning electron microscope might be beneficial (possible, if such a STEM detector is available). Therefore, the nanoparticulate material has to be applied onto a TEM support grid (with any carbon film).

Doing that has various advantages. First of all you eliminate the edge contrast in secondary-electron imaging (determines the accuracy of your particle size determination). Furthermore, almost independent of the illumination and imaging conditions you have an excellent resolution of the order of the probe diameter down to below 1 nm depending on your electron-optical system (if the projected mass density is not too high) and high contrast (depending on bright-field/dark-field mode). Due to dynamic contrast contributions you will even gain information about the inner structure of, e.g., crystalline particles like the substructure (polycrystallinity? twins?).

To comment on that in more detail more information about the material and the particle size range is needed.

Thank you Martin.

Dear Dr. Butz,

Thanks for your response.

Yes, you're right. The particles size is in range of 20-80 nm and normally nonferrous metals are as matrix.

Image J software, you can use it for size measurement.

Either use ImageJ software or digimizer. Both are free and you can google it and download. For the first time, you need to know how to use these software. So, for it you can go to youtube to watch the guideline and using method.

Thank you Geetha and Deepak.

Is it possible to change the unit of length from pixel to nm, in ImageJ software?

Generally, ImageJ automatically reads out the header of any image file, in which the magnification calibration is stored. If such a calibration is not saved in your image file, you have to calibrate your image by using /Analyze/Set scale. But therefore you need to know dimensions of any object within your image like an embedded scalebar from the annotation (or a reference image from a reference sample obtained at the same magnification).

*** Please be aware that any burned scale bar within such an image might be incorrect (especially if you want to measure distances or sizes)! So always check with at least one self-recorded reference image!***

Thank you very much Dr. Butz. Unfortunately my images are not self-recorded reference but there are embedded scalebar and after using /Analyze/Set scale I can’t change unit of length from pixel to nm or µm.

You just have to delete "pixel" and write (using your keyboard) nm/µm.

The common procedure is as follows. First, draw a line as long as your scale bar (using line selection tool). Then enter /Analyze/Set scale/. There should be already a number filled in "Distance in pixels" that indicates the length of your line. Finally, you have to enter the "Known distance" (length of your scale bar and the "Unit" using your keyboard. The software doesn't know anything about units, its just some signs that are given in the output of your measurements... ;-)

I refer to the ImageJ user guide, e.g. at http://rsbweb.nih.gov/ij/docs/guide/146-30.html.

Dear Dr. Butz,

Yes of course. Many thanks for your kind advice.

Also I have another problem. As I said, there are some agglomerated particles in my images and With respect to the applied nanoparticle size (20 nm); I want to estimate the number of particles that developed each of them according to the area of them. Although I know my images are not 3D and I can’t calculate the volume of agglomerated particles.

In imagej you can make a binary image by setting a threshold. If you use the watershed function it will break the agglomerated particles, with analyse particles you can count them individually. If you count before and after watershed you can estimate the average number of particles in a agglomerate. This does affect your measured size somewhat.

Thank you very much Dr. Verdoold. But as you mentioned, dividing agglomerated particles is estimated method (especially in binary form with several mistakes in detection) and is not accurate. I want to find more accurate method.

I think one way is calculating the area of agglomerated particles, but I don’t know how.

In imagej (or better Fiji (fjij is just imagej)) use the adaptive threshold plug-in to set a better threshold on the surface. As a complete alternative you can use MatLab with Dipimage toolbox, they have a good tutorial on counting rice grains in a photo.

Thank you Dr. Verdoold. I think this plugin is handful for images with well distinction between matrix and particles. While in some of my images, matrix is rough or many particles don’t observed clearly.

What kind of matrix and surface is it? Usually for SEM it is essential to have very good surface cleaning steps before putting the nanoparticles. (use MQ-water and piranha cleaning).

The kind of composite is aluminum matrix nano-composite. We polished and coated surface before SEM.

I find it comprehensible for the audience if the size of the particle (measured as mentioned by Bogdan and also by Benjamin) are described as a range instead of averaging the same.

Thank you Dr. Sharma.

As I said, at first step we can calculate the area of agglomerated particles. Is there any accurate method or software for such measuring?

i have used imagej but i recommend using SPIP 6 software, 

please do google it.

I can recommend the software Gwyddion. It has various image convolution functions to clean up your image and is quite quick to manually size things. It's generally designed for force microscopy but it works fine for SEM and TEM as well. Also it is free. 

Thank you James.

U can do it using MATLAB software and extract the  features

Dear Mehdi Saeidi, you can use ImageJ software. Here is link to download and set up ImageJ:

Link: https://imagej.nih.gov/ij/download.html

Video Youtube:

https://www.youtube.com/watch?v=8h3A4zB2ars&ab_channel=InnovationTechnology 

 If the nanoparticles are irregular, SEM may show a systematic bias in the measured size. The following article shows that and also proposes an enhanced accuracy method.  

Volume determination of irregularly-shaped quasi-spherical nanoparticles

http://link.springer.com/article/10.1007/s00216-016-9909-x 

http://link.springer.com/article/10.1007/s00216-016-9909-x

I use Gwyddion.

Below of SM pictures, scale is written.

Particle size is determined by identifying them, SEM machine operator to perform this task.

You need to use an image processing software. It is a little tedious. Accuracy depends on how diligent and patient you are. It is good if you use TEM to back up sizes obtained from XRD and BET 

Thank you all for this helpful information.

You can use ImageJ software to determine the average size of nanoparticles in SEM, TEM image.

Link to download ImageJ: https://imagej.nih.gov/ij/download.html

Follow this video: http://bit.ly/2sbb5B5

I recommend ImageJ

You can use matlab software with Image processing tool for measuring nanoparticles or objects in the Image.  You may need knowledge of programming.  Off course you will get better results. 

You can use image analyzer program

I recommend you Digimizer software.

http://www.digimizer.com/

thanks for answers

Image J is applicable

Software Femtoscan Program

But, as I understand, Femtoscan is for SPM images, and question was about SEM.

There is also Gwyddion software (SEM tif-format is supported):

http://gwyddion.net/features.php

Software a trial version of Origin/OriginPro.The trial version is fully functional.

www.originlab.com

I recommend you Digimizer and imageJ software.