LH/hCG-R mRNA levels during down-regulation time course. RNA was extracted from day 5 pseudopregnant control and down-regulated ovaries at 0, 6, 12, 24, 48, and 72 h after hCG injection. LH/hCG receptor levels were measured by solution hybridization assay. Total RNA (1–40 μg) was incubated with 1 × 104 cpm 35S-labeled LH/hCG receptor antisense RNA. Single stranded RNA was digested by treatment with RNase, and the double stranded RNA was recovered by trichloroacetic acid precipitation. The concentration of LH/hCG receptor mRNA was extrapolated from a standard curve, generated using increasing concentrations of the unlabeled sense strand (0–200 pg), which exhibited reproducible linearity. (Modified from Peegel, H. et al, 1994, Table 1 with permission from Endocrinology)