A) Schematic representation of the synthesis of AuMN-DTTC. The process involves sequential reduction of gold onto the parental MN, expansion of the gold seeds, and incorporation of DTTC and PEG onto the gold seeds. B) UV absorbance spectra of AuMN-DTTC, AuNP and MN. The plasmon resonance peak of AuMN-DTTC at 530 nm is an indication of the presence of gold nanostructures in AuMN-DTTC. A digital photograph of the AuMN-DTTC, AuNP and MN. The deep red color of AuMN-DTTC is a sign of gold nanoparticles in the colloidal mixture. C) R1 and R2 values of MN, AuNP and AuMN-DTTC. The R1 and R2 values of MN are comparable to those of AuMN-DTTC and significantly higher than those of AuNP (n=3; Student’s t-test p = 0.0002). D) Elemental analysis of MN, AuNP and AuMN-DTTC. The experimental AuMN-DTTC probe has iron content comparable to the parental MN and gold content comparable to the parental AuNP.

A) Transmission electron microscopy images of AuMN-DTTC suggests that the complex consists of electron-dense AuNP associated with less electron-dense dextran-coated MN. There are several gold nanostructures per probe B) High resolution-TEM (HRTEM) images show that the metallic lattices of AuNPs and dextran coated MN are located close to each other. The arrows in the magnified insets show two different metallic lattices C) EDS spectra of the dark and light colored region in the EDS map show that the nanostructures are composed of gold and iron. D) Elemental map of the probe. The gold and sulfur components co-localize with the dark colored region. The iron element co-localizes with the light-colored region of the map and is closely associated with the gold.

The absorbance spectrum of A) citrate-stabilized AuNP before and after addition 0.4 M NaCl and 1x PBS, B) AuMN-DTTC before and after addition of 0.4 M NaCl and 1x PBS, and C) AuMN-DTTC incubation in fetal bovine serum. Change in absorbance over time of D) AuNP after the addition of 0.4 M NaCl and 1x PBS (arrow shows the point of addition), E) AuMN-DTTC after the addition of 0.4 M NaCl and 1x PBS, F) AuMN-DTTC in fetal bovine serum. Unlike AuNP, the AuMN-DTTC nanoparticles remained stable long-term both in high-salt conditions and in serum.

Cysteamine induces aggregation of the probe, which can be detected by UV-Vis absorbance, relaxivity and particle size studies. Relaxivity and particle size measurements indicate that treatment with 0.4 M NaCl and 1xPBS does not induce aggregation, confirming the stability of probe. A) The absorbance spectrum of AuMN-DTTC before and 10 or 20 minutes after addition of cysteamine. Cysteamine induced aggregation of the probe is observed as a red-shift of the 530 nm peak. The horizontal arrow shows the red shift at the 530 nm peak. Vertical arrows show the decrease and increase in the 530 nm and 610 nm reading, respectively. B) Change in absorbance of AuMN-DTTC over time following the addition of cysteamine. The cysteamine-induced aggregation of the nanoparticles is accompanied by a gradual precipitation of the clusters, visible as a decrease in the 530 and 610 nm peaks. C) Change in the T2 relaxation time of a solution of AuMN-DTTC in water, 0.4 M NaCl and 1xPBS, or cysteamine. Cysteamine induces aggregation of the probe, visible as ashortening of the T2 relaxation time. Treatment with 0.4 M NaCl and 1xPBS does not produce a similar effect, confirming the stability of probe. Treatment of MN with cysteamine does not result in T2 shortening, due to the absence of the gold component. D) Particle size change of a solution of AuMN-DTTC in water, 0.4 M NaCl and 1xPBS, or cysteamine. Cysteamine induces aggregation of the probe visible as an increase in nanoparticle size. Treatment with 0.4 M NaCl and 1xPBS does not produce a similar effect, confirming the stability of probe. Treatment of MN with cysteamine does not result in particle aggregation, due to the absence of the gold component.

A) T2 weighted MR image of AuMN-DTTC and the control probes in water. The signal intensities of AuMN-DTTC and AuMN were comparable to the intensity of the parental MN and noticeably lower than those of PBS and AuNP. B) Calculated T2 values based on multiecho T2-weighted MRI. The T2 relaxation times of AuMN-DTTC and AuMN are comparable to the intensity of the parental MN and significantly lower than those of PBS and AuNP (n = 3; Student’s t-test; p < 0.05), indicating that the gold-doped probes are suitable as MRI contrast agents. C) Raman Spectra of AuMN-DTTC and control probes in water. AuMN-DTTC has a distinctive SERS signature, which is absent in the control probes.

A) A schematic of the probe injection set-up. The experimental AuMN-DTTC probe was injected in the deep right gluteal muscle. A control probe was injected in the contralateral muscle. B) in vivo T2 weighted MR image of a mouse injected intramuscularly (i.m.) with AuMN-DTTC and the control probe, AuNP. There is a notable loss of signal intensity associated with the site of the AuMN-DTTC injection, confirming the suitability of the probe as an in vivo MRI contrast agent. C) Calculated T2 values based on multiecho T2-weighted MRI. The T2 relaxation time of AuMN-DTTC was significantly lower than both non-injected muscle and muscle injected with AuNP (n = 3; Student’s t-test; p < 0.05). C) A photograph demonstrating the Raman spectroscopy experimental set-up. E) in vivo Raman spectra of a mouse injected i.m. with AuMN-DTTC and the control probe, AuMN. The in vivo Raman spectrum of muscle injected with AuMN-DTTC has a clear SERS signature, which is indistinguishable from that obtained ex vivo and in silico and absent in skin tissue and in muscle injected with the control probe.