Protocol Archive

Stereotaxic injection of viral vectors for gene knockdown experiments: The AAV vectors for knocking down the gene of interest were custom-made with Vector Biolabs using AAV-GFP-U6 vector for AAV1 packaging (Cat no, 7040, Vector Biolabs). They were then unilaterally injected into the striatum of mice as the protocol details.

Preparation of fresh paraformaldehyde for mouse perfusion and tissue fixation.

This protocol, carried out according to the manufacturer’s instructions, uses a classical ELISA experimental design with sensitive electrochemical detection to provide a measure of 𝜶-synuclein concentration in the conditioned media of cultured iPSC-derived human cells.

The protocol described here is based upon previously established methods (Kinnebrew et al., 2019; Johnson and Radhakrishnan, 2021) and has also been used successfully to label cilia in RPE cells.

Preparation of soluble and insoluble mitochondrial protein fractions from HeLa cells for mass spectrometry analysis.

The authors investigated the colocalization of GFP-LRRK2 with a lysosomal localized TMEM192-3xHA.

This protocol details the generation of A549 PPM1H BromoTag CRISPR/CAS9 knock-in cell line.

The steps of this protocol are similar for different sources of protein such as cell lysate, cytosol, or membrane samples, and EV samples. However, some specific changes depend on the protein of interest, endogenous alpha-synuclein, or type of cells, hiPSC dopamine neurons.

This protocol describes a standard procedure to generate a stable cell line for a DNAJC5 knockdown using shRNA.

This protocol describes a standard procedure to differentiate SH-SY5Y cells into dopaminergic neurons using retinoic acid.

This protocol describes how to purify mitochondria from cell culture.

This protocol describes a common procedure to perform co-immunoprecipitation.

This protocol describes the characterization of the extracellular alpha-synuclein and DNAJC5.

This protocol describes the generation of DNAJC5 KO using CRISPR/Cas9 edition.

This protocol describes the in vitro depalmitoylation assay of DNAJC5.

This protocol describes membrane and cytosol fractionation of cells expressing different DNAJC5 isoforms.

Strep pull down assay for FIP200(NTD)-TSF WT or mutants were co-transfected with GST-ATG13(363-517) and/or ULK1(MIT)-MBP.

This protocol details a protocol for the trypsin digestion of PI3KC3-C1 and subsequent analysis of digested peptides using LC-MS/MS.

This protocol details plasmid construction in general terms for the insertion of genes into the pCAG mammalian expression vector.

Protocol for preparation of mitochondrial samples for immunoblot analysis.