Undergraduate immunology lab project including opportunities for experimental design, a poster presentation, and the opportunity to partially purify antibodies from hybridoma cells to utilize in a Western blot

Abstract

Abstract Undergraduate students benefit from lab activities that more closely mirror research conditions. In this 7 week lab project, students maintain the hybridoma cell line MARC 2B7 (ATCC CRL-2509), and design a mini-research project to examine antibody production under a modified growth condition of their choice. Lab groups are responsible for making their own solutions, designing controls, and monitoring and maintaining the cell line over a two week period of time. Antibody levels are measured using a dot blot. For each phase, students are presented with a standard lab protocol accompanied by a list of suggestions and/or modifications so that the students are responsible for writing their own final protocol. For example, the dot blot protocol provided is based on chemiluminescent detection; however, the commercial secondary antibody is HRP conjugated so students are also provided the manufacturer’s information on the secondary antibody and the HRP substrate available in lab. This mirrors more real-world lab experiences, and requires students to think more deeply about the experimental design. Each lab group presents a poster on their mini-project to the class. Subsequently, lab groups use ammonium sulfate precipitation followed by a desalting column to concentrate and partially purify the antibodies from hybridoma supernatants that demonstrated positive results on the dot blots. This product is used as the primary antibody for a Western blot at the end of the experiment arc. In addition to providing a platform to discuss both monoclonal and polyclonal antibodies, and enabling students to practice experimental design, this project requires students to utilize their work from early in the semester in a productive final lab experience.