Squash procedure for protein immunolocalization in meiotic cells

Several techniques have been developed for protein immunolocalization in me iotic cells. However, most of them include treatments that lead to cell dis ruption and are only suitable for prophase-l cells. We describe a novel squ ash procedure of cell preparation for protein immunolabelling of different meiotic stages. This procedure is an alternative to both cryosectioning and whole spreading procedures. We present results obtained in mouse spermatoc ytes with three different antibodies: the MPM-2 mAb against mitotic phospho epitopes, an anticentromere serum and a polyclonal serum against the SCPB p rotein of the axial elements and lateral elements of the synaptonemal compl ex. The procedure was tested for single and double immunolabelling. With th is technique a large number of cells at different meiotic stages can be ana lysed. Cell stages are easily identified and cell and chromosome structures are preserved. Thus, it allows the study of chromosome behaviour and the r elationships between the different structural elements of the cell througho ut meiotic divisions. Our procedure is also suitable for three-dimensional (3D) analyses and proved to be reliable in a wide range of systems includin g insects and mammals. In addition, the procedure may be interesting to obt ain a rapid immunological diagnosis.