The benzene dimer is the prototypical system for the study of pi stacking, and is experimentally bound by 8–12 kJ/mol (2–3 kcal/mol) in the gas phase with a separation of 4.96 Å between the centers of mass for the T-shaped dimer. The small binding energy makes the benzene dimer difficult to study experimentally, and the dimer itself is only stable at low temperatures and is prone to cluster.

• Sinnokrot MO, Valeev EF, Sherrill CD (September 2002). “Estimates of the ab initio limit for pi-pi interactions: the benzene dimer”. Journal of the American Chemical Society. 124 (36): 10887–10893. doi:10.1021/ja025896h. PMID 12207544.

Other evidence against pi stacking comes from X-ray crystal structure determination. Perpendicular and offset parallel configurations can be observed in the crystal structures of many simple aromatic compounds.

• Sinnokrot MO, Valeev EF, Sherrill CD (September 2002). “Estimates of the ab initio limit for pi-pi interactions: the benzene dimer”. Journal of the American Chemical Society. 124 (36): 10887–10893. doi:10.1021/ja025896h. PMID 12207544.

Similar offset parallel or perpendicular geometries were observed in a survey of high-resolution x-ray protein crystal structures in the Protein Data Bank.

• Huber RG, Margreiter MA, Fuchs JE, von Grafenstein S, Tautermann CS, Liedl KR, Fox T (May 2014). “Heteroaromatic π-stacking energy landscapes”. Journal of Chemical Information and Modeling. 54 (5): 1371–1379. doi:10.1021/ci500183u. PMC 4037317. PMID 24773380.

Analysis of the aromatic amino acids phenylalanine, tyrosine, histidine, and tryptophan indicates that dimers of these side chains have many possible stabilizing interactions at distances larger than the average van der Waals radii.

• McGaughey GB, Gagné M, Rappé AK (June 1998). “pi-Stacking interactions. Alive and well in proteins”. The Journal of Biological Chemistry. 273 (25): 15458–63. doi:10.1074/jbc.273.25.15458. PMID 9624131.