What is OptiPrep?
OptiPrep™ is a sterile endotoxin test- ed solution of 60% iodixanol in water with a density of 1.32 g/ml. Iodixanol was developed as an X-ray contrast medium and has therefore been subjected to rigorous clinical testing. Iodixanol is non-ionic, non-toxic to cells and metabolically inert.
How do you dilute OptiPrep?
Solution A: Dissolve 0.85g NaCl in 50 ml water; add 30 ml of Tricine or HEPES stock solution; adjust to pH 7.4 with 1 M NaOH and make up to 100 ml.
What is a density gradient reagent?
Density Gradient Reagents In density gradient centrifugation, the reagent is a product used to assist in isolation or separation of the cells. Not only can these products speed up the process, they can also increase the purity and throughput.
How do I make an OptiPrep gradient?
Place Optiprep in the sterile 50 ml falcon tube….Do not use any needles longer than ½”.
- Wipe the sides of the tube wall with ethanol.
- Place a piece of scotch tape over the hole in the tube to prevent the gradient from leaking out.
- Place the sample into a 1.5 ml centrifuge tube and store at 4˚C until ready to use.
How do I make an Optiprep gradient?
How do density gradients work?
They work by applying a density gradient to the sample, by which the components, like EVs, move to their equilibrium density (relative to the medium), separating them based on size and mass density in the case of top-down gradients, or solely based on mass density in the case of bottom-up gradients, with denser soluble …
Is Percoll toxic?
Percoll consists of colloidal silica particles of 15–30 nm diameter (23% w/w in water), which have been coated with polyvinylpyrrolidone (PVP). The PVP coating renders the product completely non-toxic and ideal for use with biological materials.
What is the density of OptiPrep density gradient medium?
OptiPrep™ is a non-ionic iodixanol-based medium with a density of 1.320 ± 0.001 g/mL. It has been used clinically in X-ray contrast imaging and has been shown to have lower toxicity compared to ionic gradient compounds such as metrizamide.
How does iodixanol gradient work?
The iodixanol gradient in this protocol is composed of steps that separate out contaminants from an impure AAV preparation. The 15% iodixanol step has 1M NaCl to destabilize ionic interactions between macromolecules. The 40% and 25% steps are used to remove contaminants with lower densities, including empty capsids.
Why cesium chloride is used in DNA centrifugation?
Because cesium is a heavy element, a cesium salt solution is much denser than the density of most salt solutions and the cesium salt solution did not affect viruses or DNA. Cesium salt solutions had ideal properties for studying DNA.