Similar questions have been asked many times on Research Gate:
https://www.researchgate.net/post/Why_was_there_size_difference_measured_by_TEM_and_DLS
https://www.researchgate.net/post/Why_does_the_particle_size_differ_in_TEM_and_DLS_measurements
In summary-
DLS and TEM measure different properties- TEM measures the physical size of the electron dense core, whereas DLS reports the hydrodynamic size of the particle which includes a diffuse layer around the particle.
Also, TEM results represent a 2D characterisation of a 3D shape, and typically far fewer particles are analysed in TEM than DLS, where many particles are measured and anisotropy will be observed as a size larger.
DLS can be very sensitive to aggregates or dust, which may mean a larger size is incorrectly reported, and similarly, sub optimal concentration can lead to noise in the detected light scattering which can again mean large sizes are incorrectly reported.
Conversely, when analysing TEM images, it is important to consider that while nanoscale structures may be observed, this material may be fused into much larger aggregates, so the true dispersed particle size may be much larger.
Thanks for the reply Alexander. According to my understanding the difference between TEM and DLS size results is about 1/3rd. But in my case the the difference appears to be very large. My concern is that can the reason still be hydrodynamic diameter?
There are a considerable number of possible answers, though in my experience the simplest is that your sample and or scattering cell are not clean enough. The second is that the sample you are studying is bidisperse or polydisperse. What pore diameter filter did you use to clean your sample? As a further point, though it would depend on the particular correlator that you are using, is that you have set the sample width -- the width in microseconds of your first correlator channel, to be wide enough that the relaxation of the 25nm particles happens in the first channel or two.
Dear George, I used 0.45 micron filter to clean my sample. the particle was a drug encapsulated particle. Does this make a difference ?
0.45 is awfully large relative to your particles of interest. Was it a cellulosic filter...those can be unfortunate...or Nuclepore or one of the similar very uniform filters? The latter are better, though you want the holes to be appreciably largely than your particles, say 100 nm. How did you clean your scattering cells?
Dear George,
Thank you for your suggestion. I will try using a different filter. We use disposable cuvettes to measure size in our instrument. Those are cleaned with distilled water before use.
If you could post a plot of the DLS data that would be useful as well. Most likely the sample is either contaminated (then filtration will help), or aggregated (the filtration may help, possibly only for a short time, may have to optimize the sample conditions), or the sample is such low concentration that your light scattering system is only detecting noise.
First we don't even know what your 'particles' are. What is the chemistry of them? In what liquid are they? What sample preparation did you use for TEM?
OK, you show one particle with a nominal diameter of ~ 50 nm. Are you referring to 25 nm the radius as 25 nm appears as the size in your question? DLS will typically measure many billions of particles simultaneously. The phrase 'representative sampling' springs to mind. Are the rest of the particles sedimented elsewhere? Are you sure that this single particle is not a contaminant?
Take a look at:
Keys for Successful Analysis – Representative Sampling & Estimation of Standard Error Calculation
https://www.malvernpanalytical.com/en/learn/events-and-training/webinars/W190613KeysLD
Thank you so much for the insights Alan. This is drug encapsulated peptide particle suspended in water. I will try taking another image and size of these particle with improved conditions.
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