Perilipin 1, unlike the other perilipins, is thought to be restricted to the fat droplet. We reassessed its cellular distribution using the fat droplet marker CGI-58 in OP9 and 3T3-L1 adipocyte lines and in brown adipose tissue (BAT). As expected, we found perilipin 1 in the fat droplet-enriched floating fraction from centrifuged adipocyte or BAT homogenates. However, about half of perilipin 1 was suspended in the cytosol/infranate or pelleted with cellular membranes. In these fractionations, most of the fat droplet-associated protein CGI-58 was in the floating fraction. In BAT and OP9 adipocytes about a third of perilipin 1 pellets, compared with a much smaller fraction of CGI-58. Co-imaging perilipin 1 and smooth endoplasmic reticulum (ER) markers reveals both ER and fat droplet associated perilipin 1 in OP9 adipocytes. Consistent with these observations, perilipin 1 overexpressed in COS7 cells mostly fractionates with cellular membranes and imaging shows it on the ER. In 3T3-L1 adipocytes almost half of perilipin 1 floats, half is suspended as infranate and small amounts pellet. Finally, driving rapid fat droplet synthesis in OP9 adipocytes increases the intensity of perilipin 1 on fat droplets, while decreasing non-fat droplet immunolabeling. Confirming the morphological findings, fractionation shows perilipin 1 moving from the pelleted to the floated fractions. In conclusion, this study documents an expanded intracellular distribution for perilipin 1 and its movement from ER to fat droplet during lipid synthesis.