Profiling the Glycosignature of Commercial Pepsin-Soluble Type I Collagen Preparations: A MultiModal Lectin Strategy...

Type I collagen (Col-I) is widely used in biomaterials and cell biology, yet the glycosignature of commercial Col-I preparations remains poorly characterized. Here, we established an integrated lectin-based workflow to profile the glycan landscape of commercially sourced bovine pepsin-soluble Col-I as supplied for biomaterial use. Using a panel of 15 biotinylated and fluorescent lectins, we combined qualitative fluorescence staining, quantitative enzyme-linked lectin sorbent assay (ELLSA), and lectin-probed western blotting to compare three production batches. The batches shared common mannose- and galactose-reactive signals but displayed marked heterogeneity in low-abundance fucosylated and sialylated epitopes. Haptenic sugar inhibition and targeted digestion with α(1,6)-fucosidase and α(2,3)-neuraminidase confirmed lectin specificity and showed that some lectin-reactive motifs were partially masked in native films. In light of the known restriction of classical Type I collagen glycosylation to galactosyl- and glucosylgalactosyl-hydroxylysine, together with physicochemical evidence consistent with residual non-collagenous components, the complex fucosylated and sialylated signals detected here most likely arise predominantly from co-purified collagen-associated extracellular-matrix glycoproteins rather than from the collagen α-chains alone. Functionally, all Col-I-coated surfaces strongly promoted MDA-MB-231 cell adhesion, while glycosidase treatment produced batch-dependent visual trends in wound-closure kinetics without significant differences at matched time points. Overall, this work identifies a hidden source of batch variability in commercial collagen biomaterials and positions lectin profiling as an accessible quality-control tool for characterizing the full glycoprotein composition of collagen raw materials.