Elsevier

IBRO Reports

Volume 1, December 2016, Pages 54-60
IBRO Reports

A standardized and automated method of perineuronal net analysis using Wisteria floribunda agglutinin staining intensity

https://doi.org/10.1016/j.ibror.2016.10.001Get rights and content
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open access

Highlights

  • PNN intensity quantification varies across the field.

  • The “ROI” method decreases variability among researchers.

  • The “ROI” method can be automated.

Abstract

Perineuronal nets (PNNs) are aggregations of extracellular matrix molecules that are critical for plasticity. Their altered development or changes during adulthood appear to contribute to a wide range of diseases/disorders of the brain. An increasing number of studies examining the contribution of PNN to various behaviors and types of plasticity have analyzed the fluorescence intensity of Wisteria floribunda agglutinin (WFA) as an indirect measure of the maturity of PNNs, with brighter WFA staining corresponding to a more mature PNN and dim WFA staining corresponding to an immature PNN. However, a clearly-defined and unified method for assessing the intensity of PNNs is critical to allow us to make comparisons across studies and to advance our understanding of how PNN plasticity contributes to normal brain function and brain disease states. Here we examined methods of PNN intensity quantification and demonstrate that creating a region of interest around each PNN and subtracting appropriate background is a viable method for PNN intensity quantification that can be automated. This method produces less variability and bias across experiments compared to other published analyses, and this method increases reproducibility and reliability of PNN intensity measures, which is critical for comparisons across studies in this emerging field.

Keywords

Extracellular matrix
Perineuronal net
Wisteria floribunda agglutinin (WFA)
Automated image analysis

Abbreviations

au
arbitrary unit
CPu
caudate/putamen
ECM
extracellular matrix
Hip
dorsal hippocampus
NaN
not a number
OF
orbitofrontal cortex
PL
prelimbic
PFC
prefrontal cortex
PNN
perineuronal net
ROI
region of interest
SD
standard deviation
SEM
standard error of the mean
WFA
Wisteria floribunda agglutinin

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