Biochemistry 3381A Lecture Notes - Lecture 20: Fluorescence Microscope, Differential Centrifugation, Osmotic Shock
20 May 2020
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Document Summary
Check for enzyme activity to check where the protein is. Because so much of it, the results will be representative. Two types of technique: subcellular fractionation a disruptive technique, microscopy a non-disruptive technique. Homogenised in a buffer solution that is isotonic to stop osmotic damage. Mechanisms include grinding, mincing, chopping, pressure changes, osmotic shock, freeze-thawing, and ultra-sound. The samples are then kept cold to prevent enzymatic damage. It is the formation of homogenous mass of cells. It involves grinding of cells in a suitable medium in the presence of certain enzymes with correct ph, ionic composition, and temperature. For example, pectinase which digests middle lamella among plant cells. Transfer supernatant each time, leaving the pellet at the bottom of the test tube. Separated into pellets according to the following: nuclei, mitochondria, peroxisomes and lysosomes, microsomes and small vesicles, ribosomes, viruses and large macromolecules.
