A simple technique for improving chromosome spreads from clumped metaphases

A method has been described by which clumped metaphases either due to inadequate hypotonic KCl treatment or prolonged storage at 4 degrees C can be rescued. The cell pellet obtained from cell suspension following centrifugation was resuspended in freshly prepared Carnoy's fixative (1:3, acetic acid: methanol) at room temperature by vortexing. Twenty microliters of Triton X-100 at a concentration of 0.5% was added drop by drop while vortexing. Three changes with fixative containing 0.5% Triton X-100 were optimal for obtaining good metaphase spreads with complete removal of the cytoplasmic background. The advantage of this technique is that important patients' samples having clumped metaphases otherwise not useful for G-banding can be rescued and karyotyped by this method.