Protocol Archive

This protocol details the cryosectioning of the mouse brain.  

This protocol details about plasmid-reprogramming of human fibroblasts.

Associated with the following preprint (published September 30th 2021): Progranulin deficiency results in reduced bis(monoacylglycero)phosphate (BMP) levels and gangliosidosis. bioRxiv 2021.09.30.461806; doi: https://doi.org/10.1101/2021.09.30.461806

Here, authors describe an approach for purification of early/sorting endosomes, providing a means by which to examine early aspects of the endolysosomal system (Endo-IP) and to combine this with lysosome purification using Lyso-IP. This allows one to examine the proteome, lipidome, as well as electron microscopy imaging of endosomes.

Here the authors present an immunofluorescence protocol to assess the extent of colocalization between FLAG-EEA1 and RAB5 with and without the Dynamin-1 and -2 (DNM1/2) inhibitor Dyngo4a.

At acid pH B–glucosidase hydrolyses the substrate 4–methylumbelliferyl–B–D–glucopyranoside to 4–methylumbelliferone and glucose. Adding alkaline buffer stops the enzyme reaction and causes 4–methylumbelliferone to fluoresce at a different wavelength from unhydrolyzed substrate, thereby permitting its measurement in the presence of a vast excess of unhydrolyzed substrate.

The objective of this protocol is to provide directive on how to extract lipids from plasma, cells, tissue, and purified organelles for analysis by liquid chromatography (LC)-MS. This will typically yield quantitative data for more than 200 lipids, depending on the sample type analyzed, across a range of lipid classes: phospholipids, cardiolipins, sphingolipids, di- and triacyclglycerols, and cholesterylesters.

This protocol describes the standard procedure of using collagenase to passage human pluripotent stem cells (hPSCs) on inactivated mouse embryonic fibroblasts (MEFs).

Protein purification protocol for tag-less LRRK2RCKW as done by Leschziner and Reck-Peterson Labs. Same protocol can be used to purify LRRK1RCKW as well. Original protocol by David Snead. Modified by Yu Xuan Lin and Mariusz Matyszewski for publication.

This collection contains protocols which describe the standard procedure of culturing human pluripotent stem cells (hPSCs) on inactivated mouse embryonic fibroblasts (MEFs).

This protocol describes the thawing of mouse embryonic fibroblasts (MEFs) as feeder cells for human pluripotent stem cell (hPSC) culture.

This protocol describes the standard procedure of thawing human pluripotent stem cells (hPSCs) on inactivated mouse embryonic fibroblasts (MEFs).

This protocol describes the procedure of thawing feeder-free human pluripotent stem cells (hPSCs) using mTeSR-plus or StemFlex.

This protocol describes a standard procedure for subcloning of genotype-confirmed human pluripotent stem cells (hPSCs).

Mouse transcardiac perfusion protocol and storage.

The intent of this standard operating procedure (SOP) is to describe procedures for mouse stereotaxic intracranial injection surgery.

This collection describes a standard procedure for isolating single human pluripotent stem cell (hPSC) clones in a high-throughput way. This collection follows nucleofection of hPSCs as described in detail in the collection “Nucleofection (Amaxa) and electroporation (Biorad) of hPSCs.” dx.doi.org/10.17504/protocols.io.b4qnqvve

This protocol describes the experimental procedure used to measure single cell survival rates post nucleofection of human pluripotent stem cells (hPSCs).

This protocol describes a standard procedure for seeding nucleofected human pluripotent stem cells (hPSCs) in 96-well plates using limited dilution. This protocol follows nucleofection of hPSCs as described in detail in the collection “Nucleofection (Amaxa) and electroporation (Biorad) of hPSCs.” dx.doi.org/10.17504/protocols.io.b4qnqvve

This protocol describes the standard procedure preparing cell pellets from human pluripotent stem cells (hPSCs) cultured on MEFs for outsourced aCGH karyotyping.