Combined total internal reflection AF spectral-imaging and Raman spectroscopy for fast assessment of surgical margins during breast cancer surgery

Abstract

The standard treatment for breast cancer is surgical removal mainly through breast-conserving surgery (BCS). We developed a new technique based on auto-fluorescence (AF) spectral imaging and Raman spectroscopy for fast intraoperative assessment of excision margins in BCS. A new wide-field AF imaging unit based on total internal reflection (TIR) was combined with a Raman spectroscopy microscope equipped with a 785 nm laser. The wavelength of the AF excitation was optimized to 365 nm in order to maximize the discrimination of adipose tissue. This approach allows for the non-adipose regions of tissue, which are at a higher risk of containing a tumor, to be targeted more efficiently by the Raman spectroscopy measurements. The integrated TIR-AF-Raman was tested on small tissue samples as well as fresh wide local excisions, delivering the analysis of the entire cruciate surface of BCS specimens (5.1 × 7.6 cm 2 ) in less than 45 minutes and also providing information regarding the location of the tumor in the specimen. Full automation of the instrument and selection of a faster translation stage would allow for the measurement of BCS specimens within an intraoperative time scale (20 minutes). This study demonstrates that the TIR-AF Raman microscope represents a feasible step towards the development of a technique for intraoperative assessment of large WLE within intraoperative timescales.

A New Look into Cancer—A Review on the Contribution of Vibrational Spectroscopy on Early Diagnosis and Surgery Guidance

“…Furthermore, invasive carcinoma was discriminated from DCIS on 1 mm 2 tissue surfaces. Autofluorescence imaging in total internal reflection (TIR-AF) mode was combined with Raman microspectroscopy by Lizio and co-workers, in order to assess the surgical margins of fresh local excisions of large dimensions (up to 10 × 10 cm 2 ) obtained from BCS and previously diagnosed as invasive carcinomas ( Figure 5 ) [ 109 ]. The authors were able to locate the tumours in the resected specimens in 45 min.…”

“… Combined TIR-AF and Raman spectroscopy analysis of a human specimen containing a phyllodes tumour. ( a ) optical image; ( b ) ratiometric TIR-AF image; ( c ) H&E-stained tissue section; ( d ) threshold image (0.8 < T < 2), showing the sampling locations for the Raman acquisition, dotted in yellow; ( e ) average Raman spectra of stroma (blue), tumour (red) and adipose tissue (green) and corresponding standard deviation (shaded in grey) [ 109 ]. …”

The complementary value of intraoperative fluorescence imaging and Raman spectroscopy for cancer surgery: combining the incompatibles

“…The combination of other imaging modalities with selective sampling of suspicious areas by RS as a powerful labelfree analytical technique has also been explored [128,129]. Such approaches include MRI, second harmonic generation, OCT, AFI, total internal reflection fluorescence, or quantitative phase microscopy [130][131][132][133][134][135][136][137][138]. In general, the basic concept behind these multimodal approaches is to combine the high sensitivity of an imaging modality (i.e.…”

“…Another study combined fingerprint RS with wide-field AFI spectral imaging for intraoperative margin assessment in breast-conserving surgery [ 130 ]. AFI was employed to effectively ‘screen out’ the adipose tissue, enabling RS to more efficiently target the non-adipose tissue regions that are at a higher risk of malignancy, with superior diagnostic value as a result.…”

Selective-sampling Raman imaging techniques for ex vivo assessment of surgical margins in cancer surgery

“…Recently, Lizio et al 83 proposed the use of auto-fluorescence spectral imaging in a total internal reflection (TIR) imaging configuration in order to increase the efficiency of autofluorescence imaging and reduce the number of Raman spectra required for the diagnosis of large breast tissue specimens. The main aim was to improve the identification of adipose tissue by using TIR auto-fluorescence imaging so to reserve the Raman spectroscopy measurements to only non-adipose tissue areas.…”