The turkey sperm glycocalyx is known to contain residues of sialic acid, alpha-mannose/alpha-glucose, alpha- and beta-galactose, alpha-fucose, alpha- and beta-N-acetyl-galactosamine, monomers and dimers of N-acetyl-glucosamine, and N-acetyl-lactosamine. Potential changes in these carbohydrates during in vitro semen storage at 4°C were evaluated using males of both high and low sperm mobility phenotypes. Changes in carbohydrate residues were quantified by flow cytometry analysis using a battery of 14 FITC-labeled lectins in combination with control (sialylated) or neuraminidase-treated (non-sialylated) sperm. Sperm were evaluated at 0, 2, 4, 8, 12 and 24 h of storage. For control sperm, 4 different patterns of lectin binding were observed over time: 1) increased mean fluorescence intensity (MnFI) at 2h (Griffonia simplicifolia Lectin-I [GS-I)] and 8h (Ricinus communis Lectin-I [RCA-I]) that remained elevated during storage; 2) increased MnFI at specific time-points (Limax flavus Lectin [LFA], 2h; Artocarpus integrifolia Lectin [Jacalin], Succinyl Triticum vulgare Lectin [sWGA], 8h; Galanthus nivalis Lectin [GNA], 12h) followed by decreasing MnFI during the remainder of the 24h storage period; 3) increased MnFI only at the 24h time-point (Lotus Tetragonolobus Lectin [Lotus], Arachis hypogaea Lectin [PNA]); and 4) no changes in MnFI during the 24h storage period (Erythrina cristagalli Lectin [ECA], Griffonia simplicifolia Lectin-II [GS-II], Pisum sativum Lectin [PSA], Glycine max Lectin [SBA], Wisteria floribunda Lectin [WFA]). For non-sialylated sperm, increased binding of ECA, GS-II, SBA and WFA were observed at variable time-points; only Canavalia ensiformis Lectin (Con A) and PSA remained unchanged during storage. Differences between mobility phenotypes existed for lectins Con A, GS-II, LFA, PSA, SBA, and sWGA, with sperm from low mobility males exhibiting higher MnFI than high mobility males throughout 24 h of storage. We conclude that the observed increases in lectin binding during semen storage indicate an augmentation of non-sialylated terminal residues which could alter sperm antigenicity and negatively impact fertility. Further, spermatozoa from low mobility males may have higher antigenicity even before semen storage. Other possible functional implications are discussed.