Spontaneous Raman microscopy reveals the chemical composition of a sample in a label-free and non-invasive fashion by directly measuring the vibrational spectra of molecules. However, its extremely low cross section prevents its application to fast imaging. Stimulated Raman scattering (SRS) amplifies the signal by several orders of magnitude thanks to the coherent nature of the nonlinear process, thus unlocking high-speed microscopy applications that provide analytical information to elucidate biochemical mechanisms with subcellular resolution. Nevertheless, in its standard implementation, narrowband SRS provides images at only one frequency at a time, which is not sufficient to distinguish constituents with overlapping Raman bands. Here, we report a broadband SRS microscope equipped with a home-built multichannel lock-in amplifier simultaneously measuring the SRS signal at 32 frequencies with integration time down to 44 mu s, allowing for detailed, high spatial resolution mapping of spectrally congested samples. We demonstrate the capability of our microscope to differentiate the chemical constituents of heterogeneous samples by measuring the relative concentrations of different fatty acids in cultured hepatocytes at the single lipid droplet level and by differentiating tumor from peritumoral tissue in a preclinical mouse model of fibrosarcoma. (c) 2022 Author(s). All article content, except where otherwise noted, is licensed under a Creative Commons Attribution (CC BY) license(http://creativecommons.org/licenses/by/4.0/).
Broadband stimulated Raman imaging based on multi-channel lock-in detection for spectral histopathology / De la Cadena, Alejandro; Vernuccio, Federico; Ragni, Andrea; Sciortino, Giuseppe; Vanna, Renzo; Ferrante, Carino; Pediconi, Natalia; Valensise, Carlo; Genchi, Luca; Laptenok, Sergey P.; Doni, Andrea; Erreni, Marco; Scopigno, Tullio; Liberale, Carlo; Ferrari, Giorgio; Sampietro, Marco; Cerullo, Giulio; Polli, Dario. - In: APL PHOTONICS. - ISSN 2378-0967. - 7:7(2022). [10.1063/5.0093946]
Broadband stimulated Raman imaging based on multi-channel lock-in detection for spectral histopathology
Carino Ferrante;Natalia Pediconi;Carlo Valensise;Tullio Scopigno;Giorgio Ferrari;Giulio Cerullo;
2022
Abstract
Spontaneous Raman microscopy reveals the chemical composition of a sample in a label-free and non-invasive fashion by directly measuring the vibrational spectra of molecules. However, its extremely low cross section prevents its application to fast imaging. Stimulated Raman scattering (SRS) amplifies the signal by several orders of magnitude thanks to the coherent nature of the nonlinear process, thus unlocking high-speed microscopy applications that provide analytical information to elucidate biochemical mechanisms with subcellular resolution. Nevertheless, in its standard implementation, narrowband SRS provides images at only one frequency at a time, which is not sufficient to distinguish constituents with overlapping Raman bands. Here, we report a broadband SRS microscope equipped with a home-built multichannel lock-in amplifier simultaneously measuring the SRS signal at 32 frequencies with integration time down to 44 mu s, allowing for detailed, high spatial resolution mapping of spectrally congested samples. We demonstrate the capability of our microscope to differentiate the chemical constituents of heterogeneous samples by measuring the relative concentrations of different fatty acids in cultured hepatocytes at the single lipid droplet level and by differentiating tumor from peritumoral tissue in a preclinical mouse model of fibrosarcoma. (c) 2022 Author(s). All article content, except where otherwise noted, is licensed under a Creative Commons Attribution (CC BY) license(http://creativecommons.org/licenses/by/4.0/).File | Dimensione | Formato | |
---|---|---|---|
DelaCadena_Broadband_2022.pdf
solo gestori archivio
Tipologia:
Versione editoriale (versione pubblicata con il layout dell'editore)
Licenza:
Tutti i diritti riservati (All rights reserved)
Dimensione
11.34 MB
Formato
Adobe PDF
|
11.34 MB | Adobe PDF | Contatta l'autore |
I documenti in IRIS sono protetti da copyright e tutti i diritti sono riservati, salvo diversa indicazione.