Purification of lipid rafts from cells and tissues (detergent-free method)

If neither of these devices is available then a continuous gradient may be prepared from diffusion of a discontinuous gradient. Equal volumes of Solution B and 5%, 10%, 15% and 20% iodixanol would be suitable. 

For more information on gradient construction see Application Sheet S03. In the nondetergent preparation of rafts from brush border membranes [9] and neuroblastoma cells [10] the gradients were discontinuous rather than continuous. 

Formation of discontinuous gradients and underlayering the gradient with the sample are best achieved using a syringe with metal cannula (i.d. approx 0.8 mm). Metal filling cannulas can be obtained from most surgical instrument supply companies. Make sure that the tubes are filled in accordance with the manufacturer’s instructions.