Spaceflight is known to affect immune cell populations. In particular, splenic B-cell numbers decrease during spaceflight and in ground-based physiological models. Although antibody isotype changes have been assessed during and after spaceflight, an extensive characterization of the impact of spaceflight on antibody composition has not been conducted in mice. Next Generation Sequencing and bioinformatic tools are now available to assess antibody repertoires. We can now identify immunoglobulin gene- segment usage, junctional regions, and modifications that contribute to specificity and diversity. Due to limitations on the International Space Station, alternate sample collection and storage methods must be employed. Our group compared Illumina MiSeq sequencing data from multiple sample preparation methods in normal C57Bl/6J mice to validate that sample preparation and storage would not bias the outcome of antibody repertoire characterization. In this report, we also compared sequencing techniques and a bioinformatic workflow on the data output when we assessed the IgH and Igκ variable gene usage. Our bioinformatic workflow has been optimized for Illumina HiSeq and MiSeq datasets, and is designed specifically to reduce bias, capture the most information from Ig sequences, and produce a data set that provides other data mining options.

We thank Ms. Melissa Gulley for her help in the lab and Mr. Ricky J. Rettig for his help and expertise in Microsoft Excel. We also thank Drs. Ruth Globus and Sungshin Choi at NASA Ames for their help with RR1 data and Dr. Alina Akhunova, Director of the Kansas State University Integrated Genomics Facility, for her help, dedication, and sequencing expertise.