Fragile X expression increased by low cell-culture density.

The effect of cell density on expression of fra(X) was studied. A lymphoblast cell line from one fra(X) individual and whole blood from another individual was tested at various cell densities using RPMI-1640 with FUdR (0.1 microM) 24 hrs before harvest. Densities from 0.25 X 10(5) to 2 X 10(6) cells/ml were tested. Chromosomes were G-banded and analyzed for fra(X) frequency. Increased density caused fra(X) frequency to decline in lymphoblasts and whole blood. In the established line low density fra(X) frequency was 51.2% and decreased to 6.5% at the high density. In blood fra(X) frequency was 34.7% at low density and decreased to 18% fra(X) in high density. We suspect that decay of the FUdR effect may explain the results. Our results suggest that to maximize fra(X) frequency, cultures should be inititated at low density. This may be important in analysis of low-percentage fra(X) patients, nonexpressing female carriers, and obligate nonpenetrant males.