Analysis of micronized powders by Optical Microscopy offers the immense advantage that individual particles can be seen and measured. Critical factors such as sample preparation, micrsocope illumimation and resolution and also camera resolution must be optimal in order to simply view the particles accurately.
Once these conditions are met and a proper live image of the sample is achieved, powerful image analysis functions must be used in order to obtain a precise binary image of the objects to measure. This is crucial particularly at high magnifications such as 500X and 1000X and if the size distribution of the particles is wide because the largest and smallest particles lie in different focal planes.
In such conditions a multi-layer image reconstruction is necessary; without it some objects would be missing depending on focus; small objects would be missing when focusing on the large ones and large objects would be unclear when focusing on the small ones; in both cases intermediate objects would have blurry edges.
Figure A: Reconstructed image with large and small particles in focus.
Figure B: Grey Threshold on reconstructed image. Large and small particles are in focus.
Once accurate representation of the particles is achieved, relevant measures are performed on the individual objects and the results can be validated and artifacts removed (if any) prior to results output, since all the positions of the objects are remembered. This method allows for reproducibility and traceability of statistical results for detailed shape and size information of micronized powders.
Figure A: Micronized powder at 1000X. Small particles are in focus at lowest focal plane.
Figure B: Micronized powder at 1000X. Grey Level Threshold on lowest focal plane.
Figure C: Micronized powder at 1000X. Large particles are in focus at highest focal plane.
Figure D: Micronized powder at 1000X. Grey Level Threshold on highest focal plane.