A computer-implemented method and system perform efficient and accurate fiber morphology analysis of fiber-containing biological samples. The images of objects comprising fiber and/or branching structure are obtained by camera and/or other imaging devices. Initial tracing seeds are obtained by variable modules. The adaptive threshold method is utilized for identifying tracing seeds candidates by local Principal Component Analysis (PCA) calculation and sorting out the seeds with low and high score. Each single fiber segment of interest is traced repeatedly utilizing PCA fitting calculation, and individual fiber segments are assembled by performing crossover matching calculation.