A method for automatically altering an imaging area in microscopic imaging of a biological sample. In the method, a sample outline is differentiated from surrounding tissues by means of endogenous or exogenous markers; an initial sample imaging area is set; optical microscopic imaging is performed on a sample surface layer, wherein the imaging area is larger than an area to be imaged of the sample; an actual sample area is calculated by an outline identification algorithm using an imaging result of the sample surface layer and is set as an imaging area of next layer; optical microscopic imaging is performed on a sample to be imaged of the next layer according to the set imaging area, wherein the imaging area covers the area to be imaged of the sample and no redundant imaging is performed; and the above steps are repeated until a data acquisition task is completed.