Protocol Archive

This protocol details Cryo-EM grid preparation.

This protocol describes routine cell culture maintenance as described in Adriaenssens & Nguyen et al., 2023 (BioRxiv).

This protocol describes plasmid construction as described in Adriaenssens & Nguyen et al., 2023 (BioRxiv).

Protocol for performing an immunofluorescence assay with fixed HeLa cells.

Protocol for qRT-PCR sample preparation for analysis using a RotorGeneQ machine (Qiagen).

Protocol for analyzing oxygen consumption of isolated mitochondria using the Seahorse XFe96 analyzer.

Protocol for analysis of cellular ATP levels using the Promega Mitochondrial ToxGlo Kit.

Preparation of samples for measuring mitophagy levels using mtKeima reporter by fluorescence activated cell sorting (FACS).

Protocol for generating solubility calculations for mitochondrial proteins from starting 'soluble' and 'insoluble' fraction mass spectrometry data, using MaxQuant and Perseus software pipelines.

This protocol uses the Pierce™ High pH Reversed-Phase Peptide Fractionation Kit (Thermo Fisher, Cat# 84868).

TMT labeling.

Preparation of soluble and insoluble mitochondrial protein fractions from HeLa cells for immunoblotting.

Mitochondrial isolation and quantification from HeLa cells.

Protocol for performing an SDS-PAGE Western blot analysis using the BioRad Criterion Blotter system.

Protocol for preparation of HeLa cell lysates for immunoblot analysis.

Induction of starvation stress using EBSS in HeLa cells.

Mitophagy induction in HeLa cells using Difereprone (DFP).

Mitophagy induction in HeLa cells using Oligomycin/Antimycin A.

Protocol for generation and precipitation of retrovirus, and infection of HeLa cells to generate stable cell lines.

Salivary collection participant instructions using OMNIgene ORAL OM 501 device.