Optical coherence tomography (OCT) is a well-established medical imaging system that is frequently used to investigate depth-resolved tissue structure at high speed and resolution [1
]. Traditionally, OCT relies on the intrinsic scattering characteristics of a sample to create contrast [2
]. Kuranov et al.
were able to measure blood oxygen saturation level with dual- wavelength photothermal OCT using hemoglobin as the contrast agent [3
]. In comparison, Robles et al imaged the absorption of hemoglobin directly using a novel spectroscopic OCT scheme while also using spectral features to distinguish oxy- and deoxy-hemoglobin [5
]. However, exogenous contrast agents can provide unique sources of information and recently, there has been interest in advancing molecular contrast agents for OCT such as quantum dots [7
], near-infrared dyes [8
], and nanoparticles [9
Here, we examine the use of nanoparticles as exogenous contrast agents in phase sensitive OCT. Some of the advantages of using nanoparticles include their high degree of biocompatibility and the absence of photobleaching. Nanoparticles also exhibit rapid thermal responses to optical excitation, leading to a readily detectable phase signature. The plasmon resonance of the nanoparticles is highly wavelength specific, which allows multiplexing multiple probes with distinct excitation spectra [11
]. This direction has been examined previously. Adler et al.
have demonstrated that phase-sensitive OCT can be used to detect photothermal absorption by gold nanoparticles [12
]. Oldenberg et al.
showed that by varying the size and shape of metallic nanoparticles, the resonant optical frequency can be precisely tuned [13
]. Absorption at the plasmon resonance frequency causes highly localized temperature gradients around the nanoparticles, which in turn induces slight changes in the medium’s refractive index [14
]. These changes in refractive index can be detected using phase sensitive OCT with milliradian sensitivity.
Skala et al.
have demonstrated molecular imaging with photothermal OCT by tagging the epidermal growth factor receptors (EGFR) in live cells with gold nanoparticles [15
]. When cells containing both nanoparticles and high levels of EGFR were compared to cells with either low levels of EGFR or absence of nanoparticles, a three-fold increase in photothermal signal was observed [16
]. Paranjape et al.
have demonstrated detection of a single nanorose species in rabbit arteries [17
]. In addition, Wang et al.
have used nanoparticle species with dual- wavelength multifrequency photothermal wave imaging as an add-on modality to optical coherence tomography applied to detect macrophage and lipid in atherosclerotic plaques [18
]. It has also been shown that nanoparticles can be used as scattering contrast agents with low backscattering albedo in highly scattering tissues such as breast tissues [16
Despite the advantages of using nanoparticles as contrast agents, previous experiments have shown that bulk heating of the sample from repeated excitation can limit the signal to noise ratio (SNR) of the photothermal signal [12
]. If care is not taken, the generated heat also has the potential to damage tissue structures as in targeted photothermal ablation. Therefore, there is a need to balance bulk heating effects with modulation and detection schemes that maximize SNR in order for nanoparticles to serve as effective contrast agents for in vivo
photothermal based imaging.
In this work, we examine modulation techniques for photothermal OCT that can be used for simultaneous detection of several nanoparticle species. We demonstrate this technique by matching two excitation wavelengths at distinct temporal modulation frequencies with the plasmon resonances of different nanoparticle types to allow detection of both species at a single location. In addition, we characterize a single- pulse excitation scheme as a means to balance bulk heating of the sample with consideration of SNR. B-mode imaging of a tissue phantom with single-pulse excitation is presented in order to demonstrate the potential of using these methods for in vivo applications.