Identification of Specific Relaxin: MATERIALS AND METHODS(5)
After incubation, tissue slides were rinsed for 2 h with five changes of wash buffer (1% BSA fraction V, 0.2% fish gelatin, and 2 mM NaN3 in PBS). The tissues were then fixed for 30 min with 3% glutaraldehyde in PBS, rinsed for 30 min in PBS, and incubated for 30 min with 50 mM glycine in PBS. The tissues were then incubated for 4 h in blocking buffer 2 (1% BSA fraction V, 0.2% fish gelatin, 5% normal goat serum, and 2 mM NaN3 in PBS), and for 4 more hours in 800 ^l antibiotin IgG conjugated to 1 nm colloidal gold (BBInternational) diluted 1:25 with incubation buffer 2 (1% BSA fraction V, 0.2% fish gelatin, 1% normal goat serum, and 2 mM NaN3 in PBS). The tissues were rinsed for 2 h with 5 changes of wash buffer and postfixed with 3% glutaraldehyde in PBS for 30 min. buy birth control online
All slides were rinsed in copious amounts of double-distilled water for 30 min before silver intensification of the gold particles. Silver intensification was performed by incubating slides in silver solution (BBInternational) for 10 min at room temperature. After being rinsed in copious amounts of double-distilled water for 10 min, the tissue sections were dehydrated in an ascending series of ethanol, cleared in Clear-Rite 3 (Richard Allen, Richland, MI), and cover-slipped using mounting medium (Richard Allen).
Tags: Cells, Female, Human, Relaxin