Seven hybridoma lines which produced monoclonal antibodies against lens crystallins were established. They could detect alpha A-, alpha B-, gamma-, delta-, and several beta-crystallins of different species of animals with high avidities. Although immunohistological analysis of chicken and mouse lenses showed the typical distributions of each crystallin as have been reported so far, the ectopic expression of alpha B or delta-crystallin was observed in the brain or the kidney of chicken, respectively. Subsequently, crystallin production during the process of lentoidogenesis in cultures of chicken lens epithelial cells was examined with these antibodies. Western blot analysis revealed that beta-crystallins were not detected in cultured cells before lentoidogenesis, although all other crystallins were detected throughout the culture period. Immunostaining of cultures indicated clearly that expression of beta-crystallins was restricted to lentoid bodies. These data confirmed the lens fiber specificity of beta-crystallins as previously reported in the messenger RNA level. In addition, we found that small-size delta-crystallin (48 kDa) accumulated before the onset of lentoidogenesis. These results strongly suggest that the differentiated state of lens cells in vitro could be classified by examination of the expression pattern of crystallins. In addition, the anti-crystallin monoclonal antibodies produced in this study could be useful for detecting lens specificities.