Disclosed are an apparatus and a method for effectively measuring cholesterol using a small amount of blood. The apparatus for measuring cholesterol includes a cartridge where an upper case having a sample transferring membrane is movably coupled to a lower case having a sample measuring membrane; a cartridge accommodation unit configured to accommodate the cartridge therein; a memory configured to store therein setting information for sample measurement; and a controller configured to align the membranes of the upper case and the lower case with each other, by horizontally moving the lower case of the cartridge, according to the stored setting information, and configured to contact the membranes of the upper case and the lower case with each other, by vertically moving the upper case for sample transfer on each alignment position.