Effect of isotope substitution from the modulation of Thermal-Lens (TL) signals in binary liquid mixtures is explored using femtosecond pump-probe mode-mismatched Z-scan TL spectroscopy. Binary solutions of water and heavy water with pure methanol at various concentrations are used for the study, where pure methanol acts as the TL marker across the different samples. The TL signal is found to be effectively modulated by isotope substitution under the experimental conditions, indicating that TL signal depends on hydrogen bonding. (C) 2014 Elsevier B.V. All rights reserved.