Fig 2: CTC signatures for patient 1. Patient 1 was treated with dabrafenib and trametinib for 1 month and discontinued because of toxicity. Tumour progressed after cessation of treatment. a CTC signatures presented according to days of treatment. All markers fell initially corresponding to clinical response (day 40). CTC signatures changed significantly on day 48 in response to treatment with markedly reduced LNGFR level. The signatures returned to pre-treatment pattern upon cessation of treatment, and elevated intensities correlated to disease progression (day 111). b Surface marker expression profile in response to treatment. Signal distribution indicated the tightening of the surface marker expression with treatment and broadening with disease progression. c Clustering of CTC signatures in response to therapy after application of LDA on SERS signals. Data in a are mean ± s.d. with 150 measurements. Each curve in b is calculated by 150 measurements of the bulk of cells

Fig 3: Histograms showing number of quantitative analyses of immunohistochemistry performed for (a) NGFr/TrkA, (b) BDNF, (c) TrkB and (d) CALCRL in canine IVD-AF tissue sections. * p-values for statistical significance for NTG-101 as compared to PBS-injected canine IVDs.

Fig 4: Anti-CD63 functionalized EPAC specificity.The specificity was studied using EV samples released from SK-MEL-28 and MCF7 cell lines, as well as control experiments including (++) EV-free cell culture medium, (−+) without the CD63 capture antibody, and (+−) with nontarget CD45 detection antibodies on SERS nanotags. (A) The expressions of MCSP, MCAM, ErbB3, and LNGFR in SK-MEL-28 and MCF7 cells were detected by flow cytometry. (B) Representative false-color SERS spectral images, (C) average SERS spectra obtained from corresponding SERS mapping datasets, and (D) average SERS intensities at 1075 cm−1 (red, MCSP), 1375 cm−1 (blue, MCAM), 1335 cm−1 (green, ErbB3), and 1000 cm−1 (yellow, LNGFR). Data in (D) are represented as means ± standard deviation, where error bars represent standard deviation of three separate experiments. Scale bars, 10 μm.

Fig 5: CTC detection and characterisation with Raman spectroscopy. a, b Schematics of experimental workflow: the blood sample taken from a patient is first depleted of RBC and PBMCs by processing over density gradient centrifugation (Ficoll) and subsequent CD45 depletion. Remaining cells are incubated with antibody-conjugated and Raman reporter-coated gold nanoparticles (Ab-SERS labels). The sample is subsequently washed and tested with Raman spectroscopy. To characterise the CTC populations, the Raman intensities are plotted as a frequency distribution curve. This curve represents the sample’s range of expression. Higher intensity indicates the presence of more Ab-SERS labels as a result of higher marker expression levels or number of cells. Four melanoma surface marker antibodies (MCSP, MCAM, ErbB3, and LNGFR) with four specific SERS labels can be multiplexed for monitoring the CTC surface marker expression simultaneously; c CTC populations in response to treatment: the frequency distribution of each marker can signal how diverse the cell populations are in terms of surface marker expression levels. The more diverse and heterogeneous the sample population, the wider the signal distribution of the respective markers. Selection of subclones or adaptation to specific selective pressure results in on-treatment signatures with a narrowing spectrum of phenotypes, while after resistance establishment phenotypic spreading can be observed. d CTC signature in response to treatment: the relationship between the average Raman intensities of each surface maker represents the CTC signature. This signature is unique to each cell population. Shrinking of all marker intensities but retaining the relationships (CTC signature) could mean diminished cell number (tumour regression). Changing of CTC signature means the population now has a different phenotype, which could represent a treatment-resistant cell population