Construction of a system using a deep learning algorithm to count cell numbers in nanoliter wells for viable single-cell experiments.

Journal: Scientific reports
Published Date:

Abstract

For single-cell experiments, it is important to accurately count the number of viable cells in a nanoliter well. We used a deep learning-based convolutional neural network (CNN) on a large amount of digital data obtained as microscopic images. The training set consisted of 103 019 samples, each representing a microscopic grayscale image. After extensive training, the CNN was able to classify the samples into four categories, i.e., 0, 1, 2, and more than 2 cells per well, with an accuracy of 98.3% when compared to determination by two trained technicians. By analyzing the samples for which judgments were discordant, we found that the judgment by technicians was relatively correct although cell counting was often difficult by the images of discordant samples. Based on the results, the system was further enhanced by introducing a new algorithm in which the highest outputs from CNN were used, increasing the accuracy to higher than 99%. Our system was able to classify the data even from wells with a different shape. No other tested machine learning algorithm showed a performance higher than that of our system. The presented CNN system is expected to be useful for various single-cell experiments, and for high-throughput and high-content screening.

Authors

  • Takashi Kamatani
    Pulmonary Division, Department of Medicine, Keio University School of Medicine, 35 Shinanomachi Shinjuku-ku, Tokyo 160-8582, Japan.
  • Koichi Fukunaga
    Pulmonary Division, Department of Medicine, Keio University School of Medicine, 35 Shinanomachi Shinjuku-ku, Tokyo 160-8582, Japan. km-fuku@cpnet.med.keio.ac.jp.
  • Kaede Miyata
    Department of Biological Sciences, Graduate School of Science, The University of Tokyo, Bunkyo-ku, Tokyo, Japan.
  • Yoshitaka Shirasaki
    Department of Biological Sciences, Graduate School of Science, The University of Tokyo, Bunkyo-ku, Tokyo, Japan.
  • Junji Tanaka
    StaGen Co. Ltd, Taito-ku, Tokyo, Japan.
  • Rie Baba
    Pulmonary Division, Department of Medicine, Keio University School of Medicine, 35 Shinanomachi Shinjuku-ku, Tokyo 160-8582, Japan.
  • Masako Matsusaka
    Pulmonary Division, Department of Medicine, Keio University School of Medicine, 35 Shinanomachi Shinjuku-ku, Tokyo 160-8582, Japan.
  • Naoyuki Kamatani
    StaGen Co. Ltd, Taito-ku, Tokyo, Japan.
  • Kazuyo Moro
    RIKEN Center for Integrative Medical Sciences, Tsurumi-ku, Yokohama, Kanagawa, Japan.
  • Tomoko Betsuyaku
    Pulmonary Division, Department of Medicine, Keio University School of Medicine, 35 Shinanomachi Shinjuku-ku, Tokyo 160-8582, Japan.
  • Sotaro Uemura
    Department of Biological Sciences, Graduate School of Science, The University of Tokyo, Bunkyo-ku, Tokyo, Japan.