Studies in vitro and in animals have demon- strated that protamine sulfate neutralizes the anti-IIa activity of LMWH, thereby normalizing the aPTT and the thrombin time. However, protamine sulfate neutralizes a variable portion of the anti-Xa activity of LMWH.168-172 It is likely that incomplete neutraliza- tion of anti-Xa activity re ects the fact that prot- amine does not bind to LMWH fragments within the LMWH preparations that have low sulfate charge The clinical signi cance of incomplete anti-Xa neutralization of LMWH by protamine sulfate is unclear. In a small case series, protamine sulfate failed to correct clinical bleeding associated with LMWH in two of three patients,169 but there are no human studies that convincingly demonstrate or refute a bene cial effect of protamine sulfate on 2012 American College of Chest Physicians by Kimberly Henricks on February 10, 2012 chestjournal.chestpubs.org Downloaded from www.chestpubs.org CHEST / 141 / 2 / FEBRUARY, 2012 SUPPLEMENT e33S 1.3.1 Discovery of the Natural High-Af nity Pen- tasaccharide: Building on the discovery of Lindahl and associates, who isolated heparin fragments with high af nity for AT, Choay and colleagues192 and Thunberg et al193 demonstrated that the minimum heparin fragment necessary for high-af nity binding to AT consisted of a pentasaccharide. Choay and associates125,194 then isolated this high-af nity pen- tasaccharide and demonstrated that it formed an equimolar complex with AT and enhanced AT- mediated inhibition of factor Xa. In 1987, Atha and associates195 reported that both the 3-O- and 6-O- sulfated glucosamine residues within the pentasac- charide sequence were critical for its activity. These observations paved the way for the development of fondaparinux. 