Dear Svetlana:

From my point of view there are several problems that are being mixed on the subject :

1) The first problem is the interpretation of what you're watching. The two techniques although they are both electron microscopy , produce different images showing particular characteristics , just by differences in the form and manner of creating the image .

In the TEM image of what you see is almost a radiograph, in which objects or structures of high atomic number produce a dark contrast , because the electrons do not pass through as easily as in the polymer with low atomic number , although the nanoparticles are embedded , or in the interior of the polymer can be detected, just as in an X-ray radiography , we obtain a contrast due density differences detecting the a bone, although it is protected by muscles .

In contrast in SEM we observe the relief of objects, the topography of the materials or structures, but probably you can have more difficulties to detect nanoparticles embedded or included in the volume of the polymer.

2) I think , as was previously suggested , it would be desirable to obtain a chemical rather than topographic contrast , so in this case would be interesting to obtain an image of backscattered electrons (BSE ), but the problem is that the spatial resolution of that image is worse than the image of secondary electrons , so that if the SEM microscope you're using is old, or if their performance is limited , it is possible, not detected or solve particles of 15 nm.

3) You can´t compare 2 images in such different magnification ranges . The field of view of the TEM image (400nanometers ) is , at least, 25 times smaller than the SEM field of view (10 microns ) . Of course it is advisable to go to higher magnifications to try to detect such small particles .

4) Maybe to start, and if you do not have much experience in electron microscopy , it is recommended to measure the same sample by the two techniques . The TEM grid in which the sample is prepared , may also be measured by SEM .

5 ) Very importantly , the TEM microscope allows obtain images with better spatial resolution than the SEM microscope , but this should not mislead us , as the two microscopes can NOT provide completely different results if your observing the same structures. So two images of SEM and TEM should provide particle sizes quite similar, of course considering that the SEM microscope can reach sufficient resolution to resolve the nanoparticles. Therefore despite the resolution differences in both microscopes , this may not be the reason for providing such different size particles.

Have you used any sort of capping agent/stabilizer? If so, it can be transparent for TEM and at the same time it will change the picture at SEM.

Thank you for your attention to my problem. The samples for measurements were prepared separately by ion-exchange saturation and further deposition without any stabilzer. The conditions of synthesis were the same in both cases. The increase of the SEM-magnification only "stretches" previous image.

Dear Svetlana, this is the rule rather than the exception when comparing the same material using different methods/equipments; specially when dealing with nanomaterials.

I agree with previous comments. if you used stabilizing agent (as can be presumed from TEM image), you just need to switch to higher magnification in SEM. With x14000 you won't see 15-nm particles. Or they might be just embedded in your polymer

As far as I know, it is not necessary easy to see features of let us say tens of nm in conventional SEM. And for some older or lower cost equipment it may be below the ability of particular SEM. From that point of view I am afraid that You have observed different features on your samples.

I thing You should test abilities of your SEM on well-known sample (or even ”standards” for SEM testing) firstly. After that You may deal with your samples and question of nanoparticle surface treatment, agglomeration/aggregation, preparation of sample for microscopic observation, etc.

Svetlana, I think you could increase the magnification of the SEM, you are looking in TEM details obviously smaller (comparing images). Probably in TEM you are observing details in the white regions observed in SEM images. Also is possible observation of particles agglomeration in SEM, you can try also Backscattered signal. I believe is just an issue of analysis of the reults.

First test TEM of your nano-particles dispersed on lacey carbon grids and then use the same grid to test for SEM. Also avoid using Pt or Au coating in SEM test.

I working with quantom dots 655 and 525. I can see them on carbon film grids on TEM with magnification 92,000. I don't think by yours images it is quantom dots. Look on higher magnification.

In order to get an idea of the particle size it is best to first perform a DLS/NTA. While you perform the DLS keep an eye on the poly dispersity index(PDI) that way you will get an idea of the size distribution i.e. whether the particles have a uniform distribution(PDI tends to 0) or its a heterogenous population(PDI tends to 1).

That way when you you perform SEM/TEM you will know exactly what to look out for. In TEM sometimes if the grinds are too old the carbon grids have aggregates on black particles which are mostly carbon particle aggregations.

I hope this will help in your future experimental designs...

Different sizes of metal nano-particles are occur in SEM and TEM images due to its different resolutions. TEM has higher resolution than SEM, So Metal nano-particle sizes obtained from SEM techniques have larger particle size than the TEM technique.

Dear Svetlana:

From my point of view there are several problems that are being mixed on the subject :

1) The first problem is the interpretation of what you're watching. The two techniques although they are both electron microscopy , produce different images showing particular characteristics , just by differences in the form and manner of creating the image .

In the TEM image of what you see is almost a radiograph, in which objects or structures of high atomic number produce a dark contrast , because the electrons do not pass through as easily as in the polymer with low atomic number , although the nanoparticles are embedded , or in the interior of the polymer can be detected, just as in an X-ray radiography , we obtain a contrast due density differences detecting the a bone, although it is protected by muscles .

In contrast in SEM we observe the relief of objects, the topography of the materials or structures, but probably you can have more difficulties to detect nanoparticles embedded or included in the volume of the polymer.

2) I think , as was previously suggested , it would be desirable to obtain a chemical rather than topographic contrast , so in this case would be interesting to obtain an image of backscattered electrons (BSE ), but the problem is that the spatial resolution of that image is worse than the image of secondary electrons , so that if the SEM microscope you're using is old, or if their performance is limited , it is possible, not detected or solve particles of 15 nm.

3) You can´t compare 2 images in such different magnification ranges . The field of view of the TEM image (400nanometers ) is , at least, 25 times smaller than the SEM field of view (10 microns ) . Of course it is advisable to go to higher magnifications to try to detect such small particles .

4) Maybe to start, and if you do not have much experience in electron microscopy , it is recommended to measure the same sample by the two techniques . The TEM grid in which the sample is prepared , may also be measured by SEM .

5 ) Very importantly , the TEM microscope allows obtain images with better spatial resolution than the SEM microscope , but this should not mislead us , as the two microscopes can NOT provide completely different results if your observing the same structures. So two images of SEM and TEM should provide particle sizes quite similar, of course considering that the SEM microscope can reach sufficient resolution to resolve the nanoparticles. Therefore despite the resolution differences in both microscopes , this may not be the reason for providing such different size particles.

I look these photos and see on left (SEM) bright particles with size 100-200 nm (0,1 - 0,2 mkm). And on the right photo I see black particles with size ~ 100-300 nm (one of them lay just above scale). So I think that it is the same particles. At SEM image contrast formed by reflection, so particles seems bright. At TEM contrast forms by absorbtion of electrons so the same particles seems black.

At TEM image a lot of small particles are visible which does not visible in SEM due to small magnification. This produces an illusion that it ia the same particles. But it is only illusion.

Many thanks for all recommendations.

Dear Alexander, I am afraid that the dark area near scale is not a particle, perhaps it is a part of polymeric matrix which is not transparent for electrons. Although, I observed big particles/aggregates (~100 nm) with TEM on such samples before. But this is not typical. The typical picture with small particles is presented in the photo (b).

Svetlana, I heve not so much experience in TEM of polymers but this black area have flat edges and corners, close to the direct. Is it possible for polymer?

In my previous experience, i think this is due to differences in the resolution of the equipments. For SEM the resolution is quite low than that of TEM. Using SEM, i initially thought that the image is contained of a single nanoparticle. But actually, when using TEM, evident shows that it is made of an agglomeration of many NPs. So, the issue of getting different sizes of NPs is actually didn't arise.

Svetlana, to sum up many good answers given above:

I am almost 100% sure that what you are seeing as "bright spots" in the SEM image are poorly conducting pieces of your polymer/matrix. When electrons from the SEM e-gun hit them, they (electrons) stay on these pieces, since there is poor electrical contact to the ground. So next time the SEM beam comes around, it is repelled by this (by now charged) piece of material. This gives extra electron reflection -> bright spots. This is typical for insulating samples in the SEM.

You can also see that these "bright spots" have the same topology as the rest of the surface - it is all covered in such "flakes", with varying contrast from bright to dark. This strongly suggests that they are all the same (polymer), but some are better "grounded" than others.

Another argument why these are not your NPs is that the distribution of NPs on the TEM image is non-uniform (they bunch together, at places forming quasi-1D structures). On the SEM image you see none of this behavior.

Now as to how to properly determine the size of your NPs, I would say that your TEM is good enough and sufficient. HRTEM is the de-facto standard method for NP size determination in the literature.

If copper density in the polymer is sufficient, you may also try a relatively simple and widely available method of x-ray diffraction (XRD). You should get the copper lines on the theta-2theta diffractogram, and the width of each diffraction line can be converted to the average size of your NPs following the Scherrer formula. You may read more about this here: http://dx.doi.org/10.1016/j.electacta.2013.03.121 (and references therein on Scherrer analysis)

In oxide materials I am seeing this problem many time. In SEM we are seeing the aggolomorates rather than single particle. However in TEM the sample preparation process is such that you will be able to break the agglomorates and see primary particle. In SEM many times I see particle (agglomorate) of size 300-500nm and in TEM acual size of indiuvidual particle is 50-70nm.

in addition to what I saw in previous comments, I would like to add:

- the bright spots may be electrostatic discharges

- in SEM, you can only resolve around 10 nm small features if you are using a very advanced / modern SEM equipment with the capability of a magnification like 200,000 times

- you can "see" the Cu particles only if they are not covered by an organic layer (in TEM, you can "see" through the organics, with SEM, you can not), therefore, you need to etch the surface using some suitable solvent (I would not rely on that the porous polymer really allows to detect the Cu particles, I would assume or suspect they are still covered by some 100 to 1,000 nm thin polymer layer.

From my view point there is momentous problem in your work. Your nano particles have a wide size distribution and you can see different sizes in different sem or tem picture. As for my personal experiment I had synthesized nano particles that had a wide size distribution and in sem pictures I had nano particles from 50 nm to 10 micrometer. But the most imporetant thing is the average size of nano particles. You can obtain it base on your pictures.

Moreover I have a solution for you. The most precise way to find the average size of nano particles is dispering them in a solvent and take DLS test. It would give you your size distribution and the average size of your nano particles base on number of nano particles or volume of them.

TEM is the best technique for the analysis of size of the nanoparticles (especially metal) and SEM is basically used for the analysis of shape and it will not having that much resolution to analyse size.

It has to be noted that, in SEM imaging, the brightness of the spots depends on the ability of the secondary electrons to escape the surface. So, little objects or rugged objects are brighter than flat ones. Very flat substrate can show small bumps creates by nano objects. You can try ultramicrotome surfacing and high voltage acceleration in order to penetrate the surface to observe very small metallic objects embedded in polymer.

it may be possible that particles get aggregated as big sample required in SEM. but in TEM very small sample required which should be well dispersed and u can see single particle,

I would like to add this (my former comment) which should be cconsidered as well:

- the bright spots may be electrostatic discharges

- in SEM, you can only resolve around 10 nm small features if you are using a very advanced / modern SEM equipment with the capability of a magnification like 200,000 times

- you can "see" the Cu particles only if they are not covered by an organic layer (in TEM, you can "see" through the organics, with SEM, you can not), therefore, you need to etch the surface using some suitable solvent (I would not rely on that the porous polymer really allows to detect the Cu particles, I would assume or suspect they are still covered by some 100 to 1,000 nm thin polymer layer.