Measurement of polyclonal immunoglobulin synthesis using the reverse plaque technique

This unit describes the reverse hemolytic plaque assay, an effective method for measuring the number of immunoglobulin (Ig)-secreting cells present in a cell population at any particular time. Cell populations that can be assayed using the technique include peripheral blood mononuclear cells or cells from tissues such as the tonsils. The basic protocol is divided into three stages. First, protein A-sensitized sheep red blood cells (SRBC), guinea pig complement, and anti-Ig antibody are prepared. Test samples are then combined and incubated with the SRBC, complement, and antibody in appropriate chambers. Finally, the resulting plaques are scored. A support protocol describes the preparation of plaquing chambers.