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the total microglial cell size, cell body size and size In conclusion, the method for analyzing microglial
of the dendritic processes separately, the specific morphological changes as marker for microglial
characteristics of the morphological changes can be activation by semi-automatic image analysis of IBA-1
investigated. In our experiment for instance, it became stained brain sections, described in this article, provides
apparent that (without significantly changing the a novel sensitive marker for microglial activation in
number of microglia or cell size) there is a reduction of rats, which is quick and easy to perform and provides
the size of the dendritic processes. Surgery, on the other additional information about the morphological
hand, led to increased cell body size, specifically in characteristics of microglia. Although conclusions
the aged rats. Accordingly, a dystrophic deramificated about microglial activation based on morphological
morphology of microglia has been observed in old rats characteristics alone should be drawn with great
and humans, [6,28] whereas surgery was associated with caution, this novel analysis method could, combined
a more classical microglial activation. [20,29] with other markers related to microglial activation,
contribute to research in the field of neuroimmunology.
The method proposed in this article has several
limitations. Firstly, in this study we validated our ACKNOWLEDGMENTS
method on relatively small areas (0.06 mm ) in order
2
to compare it to microglial activation based on visual We acknowledge funding from the University of Groningen
characterization in those same areas. Therefore, only as this research was part of the Rosalind Franklin Fellowship
approximately 13 microglia/area were analyzed. awarded to Dr. van Leeuwen. We thank Jan Keijser, Wanda
However, we successfully explored the method on Douwenga and Folkert Postema for their technical support
substantially larger areas (data not shown), further and Kèvin Knoops for his assistance with implementing the
confirming its benefits. image analysis method for Image J. We thank Prof. EA van
der Zee, chair of the Department of Molecular neurobiology
of the University of Groningen, and Prof. E Heineman, chair
Second, although we aim for an objective marker of the Department of Surgery of the University Medical
for microglial activation, the staining threshold and Center Groningen for the use of their facilities and general
size filter have to be subjectively determined by and material support.
researchers based on the staining intensity of the
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