HIGH-THROUGHPUT SCREENING BY MULTICHANNEL GLASS-FIBER FLUOROMETRY

As a tool for screening large numbers of biological samples by means o f amplification (e.g., Q beta or PCR) we have constructed a thermocycl er that includes optionally a 96-channel or 960-channel glass fiber fl uorimeter (combined with a cooled charge-coupled-device camera). We br iefly describe the system integration of all components like liquid ha ndling, thermostats, an x,y,z robot arm, and the glass fiber fluorimet er. The integrated glass fiber fluorimeter allows sensitive on-line me asurements in 960 channels within 5 s. Two different screening procedu res were carried out. In a first experiment PCR reactions were done in the presence of the known PCR inhibitor hematin and its suppressor tr ansferrin. The system was used to titrate the suppressor with the inhi bitor hematin in order to determine the maximum inhibitor concentratio n tolerated at a given suppressor concentration. We processed 96 PCR s amples in parallel with 11 different concentration steps. In a second experiment the 960-channel glass fiber fluorimeter was used to monitor on line the amplification of the Q beta system in the presence or abs ence of an inhibitor (heparin). Since the doubling time of Q beta RNA variants is about 20 s, on-line detection is crucial for the experimen tal setup. The evolution of new RNA species adapted to high inhibitor concentrations could be proved by comparison of the fluorimetric signa l and electrophoresis. (C) 1997 American Institute of Physics.