研究現場での転写膜応用の問題点と将来的課題

抄録

The past two decades can be characterized by a revolution in our understanding of living organism. The major driving force in this revolution has been recombinant DNA techniques. The technique is characterized specially by its ease in application. The most frequently used technique is the gel electrophoresis, followed by blotting of the gel separated samples to a transfer membrane and the subsequent hybridization of the membrane with a probe.In this article, I reviewed some of the practical problems associated with the use of transfer membranes in the laboratories.
When radioactive tracers are used for detection by X-ray film, the strength of the signals above the minimun threshold of the X-ray film detection determine its usability. It was necessary to maximize the amount of the samples absorbed on the membrane, and to optimize the fixation method to get the signal strength required for satisfactory results. Among the materials used for transfer membrane, Nylon gained the popularity. When Bioimage-analyzer from Fuji Film Company was introduced, the machine dramatically changed the scope of the technique because of its high sensitivity, the accuracy in quantitation, and the flexibility for data processing by the ample power of the computer in the analyzer. The deciding factor for sensitivity in detection is now low background noise which increases the signal-to-noise ratio (S/N). Cellulose nitrate membrane and PVDF membrane are the material more preferred over Nylon in this regard.
More recently, non-radioactive detection systems are gaining popularity. The systems using Chemi-luminescence are begining to be used widely, and CCD cameras are getting better allowing the method to get the benefit of computer power. The biggest problem with this method is in the inaccuracy in quatitation due to the nature of the membrane ; e.g. the transfer membranes have thickness which is important to increase the capacity of binding samples but inhibits the light signals from within the membranes due to blockage of light by the membrane matrix. Another non-radioactive method using fluorescent probes suffers from the natural fluorescence of the membrane matrix causing the high background. This problem is additional to the light permeability problem. It is now highly desirable to develop new transfer membranes suitable for optical methods used in the non-radioactive detection systems.