golgi Protocols

Category: Cell Organelle Protocols: Subcellular Fractionation Protocols: Membrane Isolation Protocols

The protocol described in this protocol has been used principally for analyzing the Golgi, endoplasmic reticulum and trans-Golgi network but markers for other compartments (e.g. ERGIC and endosomes) have also been analyzed. Modifications either to the gradient density range or the centrifugation conditions influence the ability of the gradient to resolve multiple compartments. - [Read Fractionation of Golgi, ER, TGN and Other Membrane Compartments in Pre-Formed Iodixanol Gradients]

Category: Cell Organelle Protocols: Subcellular Fractionation Protocols: Mitochondria Isolation Protocols

Generally in iodixanol gradients the density of organelles decreases in the series: peroxisomes, mitochondria, lysosomes, ER, Golgi, although in Dictyostelium discoideum, the lysosomes are denser than the mitochondria. Iodixanol gradients can usually provide satisfactory resolution of all these membrane particles although it may be necessary to modulate either the gradient or centrifugation parameters in order to optimize a particular separation. - [Read Fractionation of Mitochondria, Lysosomes, Peroxisomes, ER and Golgi in Pre-formed Iodixanol Gradient]

Category: Yeast Protocols

Protocol for in vitro endoplasmic reticulum to golgi transport reaction in Yeast. Includes: Preparation of Membranes; One-Stage Reaction; Two-Stage Reaction using Normal Amount of Membranes; Two-Stage Reaction using Low Concentration of Membranes. - [Read In Vitro Endoplasmic Reticulum to Golgi Transport Reaction in Yeast Protocol]

Category: Yeast Protocols

Protocol for microsome high speed pellet and cytosol preparation for endoplasmic reticulum to golgi transport assay in Yeast. - [Read Microsome High Speed Pellet and Cytosol Preparation for Endoplasmic Reticulum to Golgi Transport]

Category: Cell Organelle Protocols: Microsome Protocols

Protcol relating to microsome high speed pellet and cytosol preparation for endoplasmic reticulum to golgi transport assay in yeast. Includes spheroplast formation. - [Read Microsome High Speed Pellet and Cytosol Preparation for Endoplasmic Reticulum to Golgi Transport Ass]

Category: Cell Biology and Cell Culture

Microtuble/ organelled motility assay using Golgi or ER membranes, 45 uM tubulin, rat liver cytosol, and 20x energy regeneration system. - [Read Microtubule/ organelle Motility Assays]

Category: Cell Biology and Cell Culture: In Vitro Reconstitution Protocols

Protocol includes information on: Preparation of Membranes; One-Stage Reaction; Two-Stage Reaction using Normal Amount of Membranes; Two-Stage Reaction using Low Concentration of Membranes. - [Read Protcol for In Vitro Endoplasmic Reticulum to Golgi Transport Reaction in Yeast]

Category: Cell Organelle Protocols: Subcellular Fractionation Protocols: Membrane Isolation Protocols

This protocol uses a "light mitochondrial" pellet from a mammalian liver homogenate. The gradient thus has to resolve a variety of denser components (peroxisomes, lysosomes, mitochondria) from the Golgi membranes, which have a low density in iodixanol (1.06-1.09 g/ml) [1]. The protocol is specifically tailored to the purification of Golgi membranes from this pellet and is unsuitable for the isolation or analysis of other organelles present in the light mitochondrial fraction. - [Read Purification of Golgi Membranes from a Light Mitochondrial Fraction in a Self-Generated Gradient]