Ultrafast spectroscopy studies have been key to the development of optical materials, including colloidal semiconductor nanocrystals (NCs) engineered for lighting and light-harvesting technologies. Several physical processes, which are revealed by ultrafast spectroscopy in NCs, are highly dependent on the average number of excitons created per NC, including optical gain properties and multiexciton interactions. Consequently, proper considerations regarding NC populations are necessary to avoid misinterpretations. In this paper, we present an experimental and theoretical analysis of the influence of the sample optical density (OD) at the excitation energy on the results of ultrafast spectroscopy studies in NCs. We show that the pump beam depletion caused by high ODs can drastically change the results from transient absorption (TA) experiments leading to data misinterpretations, such as the overestimation of the optical gain threshold. Based on that, we propose a robust modification on the TA technique, which allows for an OD-independent characterization, free of distortions. The modification consists of pumping the sample below its band gap energy, limiting the electronic excitation to a two-photon absorption process, resulting in an effectively zero OD for the pump beam and a uniform excitation in the direction of the beam propagation. Consequently, an undistorted TA signal is produced, allowing for precise characterization of NCs, including the gain/absorption cross section, gain coefficient, and gain threshold. Furthermore, the uniform excitation allows for higher signal-to-noise ratio, independent of the sample concentration.