Imaging nano-objects by linear and nonlinear optical absorption microscopies Mary Sajini Devadas, Tuphan Devkota, Paul Johns, Zhongming Li, Shun Shang Lo, Kuai Yu, Libai Huang 1 and Gregory V Hartland Department of Chemistry and Biochemistry, University of Notre Dame, Notre Dame, IN 46556-5670, USA E-mail: ghartlan@nd.edu Received 2 April 2015, revised 4 May 2015 Accepted for publication 18 June 2015 Published 12 August 2015 Abstract Absorption based microscopy measurements are emerging as important tools for studying nanomaterials. This review discusses the three most common techniques for performing these experiments: transient absorption microscopy, photothermal heterodyne imaging, and spatial modulation spectroscopy. The focus is on the application of these techniques to imaging and detection, using examples taken from the authors’ laboratory. The advantages and disadvantages of the three methods are discussed, with an emphasis on the unique information that can be obtained from these experiments, in comparison to conventional emission or scattering based microscopy experiments. Keywords: transient absorption microscopy, nanoparticles, imaging (Some figures may appear in colour only in the online journal) Introduction The ability to detect and analyze single molecules and nano- particles has revolutionized many areas of science [71, 80, 112, 166, 167, 175]. Some of the most important examples are in biology, where imaging at high spatial resolution has led to advances in our understanding of how molecular motors work [43, 139, 155, 176], and the kinetics of enzymes [38, 89, 174]. Typically these experiments are performed using fluorescence detection: the molecule or particle is excited at one wavelength, and its emission is detected at another, red-shifted wavelength [112, 166]. The excitation light can be removed using filters, giving a zero background signal that can be detected with high sensitivity. However, fluorescence does not work well for materials that are only weakly emissive, such as metal nanoparticles. Large metal nanoparticles (diameters greater than 20 nm) can be observed by Rayleigh scattering [136, 149, 150, 169], but small particles do not scatter strongly enough to be imaged using current state-of-the-art detectors. This is especially true for materials with resonances in the near-IR, which is an important region for biological imaging [42, 123, 168]. The shortcomings associated with fluorescence detection and Rayleigh scattering has led to the development of absorption-based microscopy techniques for single particle studies [18, 26, 40, 159]. The advantages of absorption measurements are, first, absorption can be applied to any material, even materials with small emission quantum yields. Second, absorption scales as the volume of the nano-object, rather than the volume squared for scattering [159]. This means that absorption can be used to probe much smaller objects than scattering. It is also possible to use transient absorption measurements to study dynamics in single nano- particles with a few hundred femtoseconds time resolution [20, 116, 151, 157]. In contrast, the time resolution of fluorescence measurements for single particles is typically limited to several hundred picoseconds by the response time of the detector. Gating approaches can improve the time resolution [15, 37, 57, 58], but at the expense of sensitivity. The goal of this paper is to discuss the three most commonly used absorption techniques for detecting single particles (and even single molecules). These are: transient absorption microscopy (TAM) [116, 157], photothermal heterodyne imaging (PHI) [5], and spatial modulation spectroscopy (SMS) [1]. Methods based on using a balanced detector [22, 81], interferometric detection [31, 111], ground state Nanotechnology Nanotechnology 26 (2015) 354001 (15pp) doi:10.1088/0957-4484/26/35/354001 1 Current address: Department of Chemistry, Purdue University, West Lafayette, IN 47907, USA 0957-4484/15/354001+15$33.00 © 2015 IOP Publishing Ltd Printed in the UK 1