Computational protocol: Structural heterogeneity and intersubject variability of Aβ in familial and sporadic Alzheimer’s disease

Similar protocols

Protocol publication

[…] Curcumin-, BF-188–, and FSB-labeled Aβ deposits were imaged in the spectral (Lambda) scan mode of a Leica SP8 confocal microscope using a 40× water immersion lens (1.1 NA), a 405-nm laser for fluorescent amyloid-dye excitation, and a HyD detector at 512- × 512-pixel resolution. For each field-of-view in the brain slice experiments, the optical plane was moved to the center of the z-stack volume for a given Aβ deposit, and fluorescence emission was acquired from a series of 40-image steps spanning 385- to 765-nm wavelengths using a 15-nm-wide detection window at each interval. For each field-of-view in the gel-embedded synthetic fibril experiments, confocal z-stack at multiple random positions in a well was acquired with identical spectral parameters as described above (XYλZ mode) using a 63× water immersion lens (1.2 NA). The size of the z-stack ranged from 100- to 200-µm thick depending on the density of aggregates in a given well. Raw spectral data were analyzed with Leica LASAF and NIH ImageJ software. [...] The signal-processing algorithm for the analysis of the spectra was implemented in Python using NumPy and sklearn packages. Sudden spikes in each spectrum (a result of occasional noise in the photodetector) were first removed using a five-element sliding window spline function, and all spectra were rescaled to a [0, 1] interval. To combine the spectral information from the different dyes, a new dataset of multispectral vectors was built. For a given patient and localization, a multispectral vector was composed by randomly concatenating an emission spectrum from each dye. Forty to 70 vectors were assembled for each patient. A PCA was then performed on the obtained dataset, and each vector was projected into the eigenspace defined by the two first components (covering about 60–75% of the variance). […]

Pipeline specifications

Software tools ImageJ, Numpy
Applications Miscellaneous, Laser scanning microscopy, Microscopic phenotype analysis
Organisms Homo sapiens, Mus musculus
Diseases Alzheimer Disease, Cerebral Amyloid Angiopathy