The lipid cubic phase or in meso method for crystallizing proteins. Bushings for better manual dispensing

Abstract

The lipid cubic phase or in meso method can be used to set up crystallization trials of soluble and membrane proteins. The cubic phase is noted for being a sticky and viscous mesophase. Dispensing the protein-laden mesophase by hand into wells on crystallization plates has been facilitated by the use of an inexpensive repeat dispenser. However, the assembled dispensing device is prone to damage. Specifically, the delicate plunger used to dispense the viscous mesophase by positive displacement can be bent and scarred when the locking nut that fixes the plunger to the ratchet-driven dispensing mechanism is inadvertently overtightened. A defective plunger can render the device useless as a dispensing tool. More importantly, it can lead to catastrophic loss of valuable protein and lipid due to leakage when the dispensing syringe is being recharged with fresh mesophase. This note describes two types of bushings designed to protect the plunger from mechanical damage, which provide facile and reliable dispenser performance. One is a split bushing in brass and is a highly durable solution. The other is a small sleeve made from readily available plastic tubing. While it lacks durability, the plastic sleeve is simple yet highly effective, and can be replaced as the need arises