Self-assembled monolayer (SAM) of n-alkanethiols of different chain lengths (n = 2, 3, 6, 11, 16) on a gold surface are used to immobilize antibodies which in turn bind to a antigen. The antibody and antigen used in this study have similar molecular weights i.e. ∼150 kDa. The antibody [1.5 μg cm-2] immobilized varied with the surface packing density of the SAM of carboxylic acid-terminated n-alkanethiols of different lengths. In comparison, the efficiency of antibody immobilization was lowest on the loosely packed SAM of n-alkanethiols (n ≤ 3) and the highest on the densely packed SAM of n-alkanethiols (n ≥ 11). However, increased immobilization of antibodies with increasing chain length of the n-alkanethiols [n > 11], did not result in a corresponding increase in antigen binding. An attempt to explain this phenomenon based on packing density and an orientation of the captured antibody is presented. © 2015 The Royal Society of Chemistry.