Beginning at Separation Lake Trailhead, Separation Lake Trail #3536 immediately enters the Three Sisters Wilderness. The trail travels through unique forest with many tree species, including Sugar Pine, Pacific Yew, Silver Fir, and more. Following a ridge downhill, the trail eventually crosses Separation Creek. Currently, a bridge is in place for hikers to cross on, and another wet crossing exists for stock. On the south side of Separation Creek, the trail heads west uphill through forest and swamps before reaching Separation Lake. The trail continues past Separation Lake to very steep uphill switchbacks before crossing Harvey Creek and ending at Horse Creek Trail #3514.