The present work investigates the impact of dietary-carbohydrate chain length on the binding affinity with Agaricus bisporus lectin (ABL) using sugars of varying chain length, including glucose, maltose, maltotetraose, maltohexaose, maltoheptaose and maltodecaose. Fourier transform infrared analysis (FTIR) and circular dichroism (CD) record alterations to the protein secondary structure culminating with maltohexaose binding. Binding affinity was determined via intrinsic fluorescence quenching analysis and revealed that binding strength increased with ligand molecular weight, reaching a maximum at a chain length of 6 glucose units, with a Ka of 1.93x106 M-1, before collapsing for carbohydrates of higher molecular weights, in line with the trend observed for CD. Molecular docking highlights the likely binding positions of each ligand to the ABL molecule, further arguing for the presence of stable interactions between host and guest. Multiple hydrogen bonds are formed in each case, providing stabilization of the complex in the presence of additional Van der Waals forces. Maltohexose again showed the strongest interaction with ABL, with a calculated binding energy of -32.6 kJ/mol. These findings give a deeper understanding of the impact of dietary-carbohydrate chain length in relation to their binding affinity with ABL, hence advancing its potential application as a functional ingredient in highly selective therapeutics.