Sterilization is faster, easier, and less expensive than it is for conventional glass spreaders.
An improved method of sterilizing petri-dish spreaders and a spreader design to implement the method have been developed. In comparison with the conventional methods of sterilizing petri-dish spreaders, the improved method is expected to prove easier, safer, less time-consuming, and less costly, and to require less laboratory space. This improved method could be used in microbiological investigations in microgravity (e.g., aboard the International Space Station) or in normal Earth gravity (e.g., in government and clinical laboratories and research institutions). Particularly, when used in spaceflight, the improved method will prove safer than the traditional flame sterilization method.
The prior art in the design, use, and sterilization of petri-dish spreaders is relatively straightforward. A conventional petri-dish spreader is a glass rod bent like a hockey stick. The longer straight segment of the rod is used as a handle, while the shorter straight segment is used to spread a suspension of micro-organisms with a small volume (typically 0.1-mL) evenly over the surface of the growth medium in a petri dish. After a suitable incubation time, colonies of the microbes grow on the surface of the medium. Provided that a known volume of a microbial suspension is spread evenly over the medium, an accurate count can be made, and the types of organisms in the suspension can be identified. If the suspension is not spread correctly, the growing colonies converge and become so crowded that it is hard to see the boundaries between them.
Each time a suspension of micro-organisms is spread over a different dish, a sterile glass spreader must be used. A spreader that was used on another dish can be reused if it is dipped into 95-percent ethyl alcohol and the alcohol is then ignited with a flame to sterilize the spreader. If multiple dishes are to be inoculated and insufficient time is available for sterilization of a single spreader between dishes, then multiple sterile spreaders must be prepared in advance, and considerable laboratory space must be allocated for storing them. Moreover, the use of alcohol and an open flame to sterilize spreaders introduces a fire hazard.
In the improved method, the spreader comprises a forked handle and a rake made of a coil of chromel electrical-resistance heater wire (see figure) that draws a current of 3 A when a potential of 6 V is applied. All that one need do to sterilize the rake is to place the end terminals of the wire in contact with the output terminals of a 6-V power source, which can be the secondary terminals of a 120-to-6-V step-down transformer. In this method, one does not attempt to sterilize the entire spreader; only the rake is sterilized. Tests have shown that the rake becomes heated to sterilizing temperature within only 2 to 3 seconds of heating at 6 V. After electrical heating, the rake can be cooled quickly on a damp sterile pad and is then ready to use. The sterility of the spreader in this improved method has been verified by extensive testing involving spreading on sterile media after spreading 0.1 mL of a bacterial solution on a petri dish.
The improved method has proved to be very reliable. Although it involves heat, this method, unlike the flame sterilization method, does not involve either alcohol or an open flame, and thus introduces less of a fire hazard. Because this method makes it unnecessary to use multiple presterilized glass spreaders, it reduces (relative to the flame sterilization method) the amount of storage space needed. Because sterilization and cooling before each use take only a few seconds in this method, preparation time is reduced, relative to that of the flame sterilization method. A further advantage of this method is that, because of its simplicity, little (if any) maintenance is needed.
This work was done by Duane Pierson of Johnson Space Center and Thomas C. Molina of KRUG Life Sciences. For further information contact the Johnson Technology Commercialization Office at 281-483-3809. MSC-22903