Figure 3.

Glycosylation and membrane association of prominin. (A) Membranes (50 μg protein) prepared from either the brains of 12-day-old mouse embryos (E12) or adult mouse kidneys (40,000 × g, 30 min) were dissolved in SDS, incubated in the absence (−) or presence (+) of 0.8 units of PNGase F (Boehringer Mannheim) and analyzed by immunoblotting using mAb 13A4 (1 μg/ml) followed by the ECL system (Amersham). (B) A homogenate of adult mouse kidney was fractionated into a total membrane and cytosol fraction and the total membranes were treated with carbonate at pH 11 (27), yielding pH11 membranes and the pH11 supernatant. Corresponding aliquots of these fractions were analyzed by immunoblotting using mAb 13A4.