The detergent Tween‐20 solubilized preferentially portions of the marginal regions of Spinacea oleracea L. thylakoid membranes and, thus, opened the inside of the grana to the external media. Differential centrifugation. following Tween‐20 solubilization. enabled separate fractions of grana and stromal‐exposed membranes to be isolated. Analysis of Tween‐20 solubilized material, after pelleting all membrane material by centrifugation at 100 000 g, revealed polypeptides associated with the coupling factor (CF1) particles, cytochrome b6/f and photosystem II complexes, suggesting that the marginal membranes contain these proteins. Concomitantly, the 100 000 g pellet was depleted in cytochrome b6/f and P700, determined spectroscopically, Thus. our results reveal the margin to be a distinct membrane region, which does not contain the light‐harvesting centers of photosystem II (LHC II). The implication of these results, in terms of the energetic interaction of components of granal and stromalexposed membrane regions, is discussed.