Prenatal fragile X detection using cytoplasmic and nuclear-specific monoclonal antibodies

We have been carrying out studies aimed at improving prenatal detection of the fragile X chromosome/mutation, Our current protocol requires a turnarou nd time (TAT) of several days. In an attempt to reduce the TAT, we have tur ned to the use of monoclonal antibodies (mAbs), Monoclonal antibody 1A1 (pr ovided by Dr. Mandel of INSERM) immunostaining was performed according to a modified three-step immunocytochemical procedure. We found that cytoplasmi c staining intensities, using mAb 1A1/avidin biotinylated complex/diaminobe nzidine, varied from light to heavy within each sample, with controls exhib iting a majority of heavily stained cells in both chorionic villus (CV) sam ple and amniotic fluid cultured cells, Using mAb 1A1 and a new nuclear-spec ific antibody, mAb 3F11, we found that CV cultured cells harboring the FMR1 full mutation could be distinguished from controls as early as 10 weeks of gestation, in both male and female specimens. Western blot analysis showed that the antibodies have similar staining patterns hut that mAb 3F11 has f ewer background/nonspecific bands. Our results demonstrate that it is feasi ble to detect fragile X full mutations within one day after obtaining cells from CV specimens taken as early as 10 weeks of gestation. (C) 1999 Wiley- Liss, Inc.