Determination of Size and Concentration of Gold Nanoparticles from Extinction Spectra Nikolai G. Khlebtsov* Institute of Biochemistry and Physiology of Plants and Microorganisms, Russian Academy of Sciences, 13 Prospekt Entuziastov, Saratov 410049, Russia, and Saratov State University, 83 Ulitsa Astrakhanskaya, Saratov 410026, Russia Extinction spectra of colloidal gold can be used for a simple and fast determination of the size and concentra- tion of nanoparticles. It is generally accepted that experi- mental correlations of the particle size and concentration with the plasmon resonance properties are in agreement with Mie theory simulations. Here, we discuss this point in the context of a long-term collection of published experimental data and our T-matrix simulations, which account for deviations of the particle size from ideal monodisperse spheres. These deviations result in small but quite evident disagreements between the Mie calcula- tions and the experimental calibration curves “particle size vs resonance wavelength”. We present a long-term- averaged analytical particle-size calibration and also dis- cuss the effects of the particle dielectric functions, shape and size polydispersity on simulated correlations between the extinction spectra and the average particle size, and concentration. In the past few years, gold nanoparticles (NPs) 1 have attracted interest as a novel platform for various applications to nanobio- technology 2 and nanomedicine 3 because of convenient surface bioconjugation with molecular probes and remarkable plasmon- resonant optical properties. 4 Recently published examples include applications of NPs to biosensorics, 5 genomics, 6 clinical chemis- try, 7 immunoassays, 8 optical imaging of biological cells (including cancer cell imaging with resonance scattering, 9 optical coherence tomography, 10 two-photon luminescence, 11 and photoacoustic 12 techniques), targeted drug delivery, 13 immune response enhance- ment, 14 and cancer cell photothermolysis. 15 Most of these examples are based on a combination of biological recognition (the probe molecule + the target molecule) and resonance absorption or scattering of light on frequencies corresponding to excitation of localized plasmons. 16 Moreover, molecular recogni- tion has recently been used for a DNA-programmable formation of nanoparticle methacrystals. 17 Accurate determination of the size and concentration of NPs is essential for most biomedical applications of NPs. 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