Abstract text: Plant cell walls are dynamic structures composed of interdependent polysaccharide networks that coordinate growth, development, and environmental responses. Although many genes involved in wall biosynthesis have been identified, the system-level consequences of perturbing specific biosynthetic pathways remain poorly understood. Here, we applied a Gel Permeation Chromatography–Enzyme-Linked Immunosorbent Assay (GPC-ELISA) workflow to characterize changes in molecular size distribution and relative abundance of matrix polysaccharides in Arabidopsis mutants. Mutants affecting xylan (irx14L, irx15, gux1-1, gux2-2), mannan (csla2-1), pectin (gaut10-2, gaut11-2), and wall acetylation/methylation (axy4-4, tbl2, tbl3, tbl31, tbr) were analyzed to assess cross-polysaccharide responses. Disruption of one biosynthetic pathway frequently resulted in compensatory shifts in other polysaccharide classes, highlighting extensive cross-regulation during wall assembly. GPC-ELISA profiling revealed distinct signatures in polymer size and epitope abundance across mutant backgrounds, providing a comparative framework to study wall remodeling dynamics. Our results demonstrate the potential of integrated size-fractionation and immunodetection approaches to uncover hidden structural responses within the cell wall matrix and offer new perspectives for engineering biomass composition.