Culturing, transfection and NBD-labeled lipid uptake assay in HeLa-CDC50A-OE cells
By onAssay for NBD-labeled lipid uptake in HeLa-CDC50A-OE cells includes seeding, transfection, and flow cytometry to measure flippase construct activity, enabling quantitative assessment of lipid uptake.
all protocols related to “Spermidine suppresses glial inflammation and parkinsonian abnormalities in ATP13A2 deficiency”
By onAll protocols related to "Spermidine suppresses glial inflammation and parkinsonian abnormalities in ATP13A2 deficiency"
Active Escape Protocol (Stasiak et al., 2026)
By onProtocol for Active Escape task by Stasiak et al., referenced in "Integrated Representations of Threat and Control in the Lateral Frontal Pole" (2026) manuscript.
in vivo TurboID
By onProtocol uses iBioID with Lamp1-TurboID in astrocytes to study endo-/lysosomal protein interactions in mice. AAV delivery allows selective biotinylation for proteome mapping, offering insights into astrocyte signaling in vivo.
Protein quantification by SDS-PAGE and Coomassie staining
By onThis protocol provides a step-by-step guide for quantifying protein concentration by SDS-PAGE and Coomassie staining.
Immunoblotting protocol
By onThis protocol outlines a step-by-step method for performing western blot analysis, including protein separation by gel electrophoresis, transfer to membranes, antibody incubation, and detection.
Measurement of ATP13A4 ATPase activity using ADP-Glo Max Assay
By onThis protocol describes a luminescence-based ADP-Glo Max assay (Promega) to measure the ATPase activity of ATP13A4 by detecting the rate of ATP hydrolysis.
Fluorescently labeled polyamine uptake (flow cytometry)
By onAssess polyamine uptake capacities of a specific cell line after incubation with fluorescently labeled polyamines and mean fluorescence intensitiy acquisition via flow cytometry.
Auto-(de)phosphorylation assay
By onProtocol measures ATP13A4 phospho-enzyme levels by detecting its phosphorylated intermediate on an aspartate residue. Utilizes SDS-PAGE under acidic conditions and autoradiography to detect -labeled intermediates.
RT-qPCR (cell lines)
By onThis protocol describes the use of RT-qPCR to assess msAtp13a4 mRNA expression following lentiviral knockdown in C8-D1A cells.
Fluorescently labeled polyamine uptake (confocal microscopy)
By onMethod visualizes polyamine uptake in cells using fluorescently labeled polyamines and confocal microscopy.
RT-qPCR (primary cells)
By onProtocol details qPCR method for Atp13a4 expression in astrocytes and neurons using Fast SYBR Green Master Mix in a 96-well format. Normalization to 18S gene ensures accurate quantification for reliable gene expression analysis.
Developmental milestones assessment in newborn mice (P1–P21)
By onBehavioral tests for newborn mice from P1 to P21 assess weight gain, reflexes, and movements with a focus on developmental milestones.
Astrocyte morphology assessment in astrocyte-neuron co-cultures
By onThis protocol outlines the process for analyzing astrocyte morphology in astrocyte-neuron co-cultures using immunostaining and Sholl analysis.
Multiplexed immunofluorescence and RNA-FISH
By onProtocol detects Atp13a4 mRNA in astrocytes of visual cortex using immunofluorescence and RNA-FISH. Sections from Aldh1L1-EGFP mice are stained, hybridized with probes, and imaged for spatial analysis of Atp13a4 expression in postnatal development.
Glia-Free Cortical Neuronal Feeding Schedule – Neuronal Morphology
By onThis protocol outlines the treatment of cortical neurons with astrocyte-conditioned medium (ACM) or spermidine and subsequent analysis of dendrite length.
24-Hour Home Cage Activity Monitoring in Mice
By onProtocol assesses 24-hour home cage activity in adult mice using infrared beam-break technology. Beam crossings indicate horizontal locomotion continuously throughout light-dark cycle.
Astrocyte Isolation and Transfection for Astrocytes/Neuron co-culture assay
By onProtocol for isolating rat cortical astrocytes and producing astrocyte conditioned media for synaptogenesis assays.
Morris Water Maze: Spatial Learning and Memory Assessment in Mice
By onThe Morris Water Maze task assesses spatial learning and memory in adult mice by training them to find a hidden platform using spatial cues. Probe trials are conducted weekly to evaluate memory consolidation.
Open Field Exploration: Locomotor Activity and Anxiety Assessment in Mice
By onProtocol assesses locomotor activity and anxiety in mice using an open-field arena. Path length & corner crossings indicate activity level, center entries reflect anxiety levels or conflict resolution.
