Amniotic band syndrome is typically diagnosed at or soon after birth based upon characteristic physical findings. According to some researchers, minimal diagnostic criteria consist of the detection of certain abnormalities of the arms, legs, fingers, and/or toes, i.e., ring-like constriction or amputation defects.

In some cases, the condition may be suspected before birth (prenatally), based upon the results of certain specialized imaging techniques, such as fetal ultrasonography and amniocentesis, which may reveal the characteristic defects. During fetal ultrasonography, reflected high-frequency sound waves are used to create an image of the developing fetus. During amniocentesis, a small, hollow needle is used to draw out a sample of the amniotic fluid for study. In some cases, laboratory testing may detect elevated levels of alpha-feto protein (AFP) in the amniotic fluid surrounding the fetus. Alpha-feto protein, a protein produced by the fetal liver, is excreted in urine into the amniotic fluid. Raised AFP levels may be associated with a number of abnormalities in a developing fetus (e.g., neural tube defects, esophageal atresia, etc.) or may occasionally be present when the fetus is developing normally.