The organization of endoplasmic reticulum export complexes

By using relatively thick sections corresponding to the average diameter of an ER bud and the above two criteria, we were able to avoid double counting most of the ER buds present on consecutive thin sections. To establish this point independently, we performed a separate experiment in which the total number of ER buds was counted on several stacks of serial thin-sections. By comparing the number of ER buds determined by this reconstruction approach to the number determined by counting of individual sections (indirect method), we found that overestimation by the latter technique was <10-15%. Hence, both methods could be used interchangeably. We also found that the indirect method showed a high reproducibility. In spite of the high degree of ER bud enrichment in local zones, the comparison of estimates of the total number of buds per cells between groups of as few as 35 randomly taken cells with a total surface of section of 2,500 tJ.m 2 gave a value of variance of less than 5% from one group to another. In addition, in four different experiments, the value of the total number of buds per cell was found to be the same. Because of technical simplicity, we routinely used the indirect method to estimate the average number of buds per cell.