My research activity for the master thesis has been related to biophotonics looking for a way to help people by developing innovative devices and by improving laser applications in the biomedical field. This area of photonic applications has led to the concept of “lab-on-chip”, a new class of chips that integrate different biochemical analyses that can then be processed on a very small area. According to statistical results, cancer is now the second cause of death in the world: this observation was a strong motivation for focusing my thesis work on this issue. External optical setups for lab-on-chip readout are often bulky and limit a wider use of these innovative devices. Integration of the whole readout equipment has to be pursued for increasing their exploitation in the medical field. Lab-on-chips are usually low-cost, user-friendly and in order to exploit these features it would be important to pave the way to a new portable system for cancer diagnosis. To achieve such goal, we need a laser source, a spectrometer, a suitable lab-on-chip and a probe for optical readout. The aim of this master thesis is the design of a fiberoptic-based Raman probe for cancer cells recognition in human bio-fluids. Among different optical detection methods, we decided to rely on Raman scattering being a label-free technique since chemical compounds can be identified by their unique Raman fingerprint. The investigated Raman probe is designed for the medical field since our aim was to investigate the presence of cancer cells in human bio-fluids such as urine, blood, tears, sweat and saliva. In chapter 1, I have introduced cancer as the disease of the century. I have focused my attention on blood and urine analytes and their link with cancer cells. I have introduced the concept of lab-on-chip, discussing the state-of-art of these chips and the materials used in the manufactural processes. In chapter 2, I have illustrated the reasons why Raman spectroscopy is one of the best optical detection method for lab-on-chip starting from the principles of the Raman effect. I have described own freeform reflector Raman lab-on-chip used in the laboratory and its manufacturing process. In chapter 3, I have discussed the design of a fiberoptic-based Raman probe, starting from the concept of optical probe. I have then shown the so called “coaxial probe design”. This layout uses a single mode optical fiber to guide the excitation signal over the detection area of the chip and one multimode fiber to collect the excited Raman scattering. A dichroic mirror separates the excitation light from the detected signal while lens, mirror and optical filters are used to enhance system throughput and signal-to-noise ratio. For the ray-tracing simulations, I relied on ASAP (Advanced Systems Analysis Program) to get the optimum layout and Matlab for the Raman scattering simulation in the detection area. The numerical results have been important to define the geometry of the optimum probe, which has a length of 138.9 mm and a width of 13.3 mm while the best working distance is 5 mm. I have imposed to ensure a tolerance of ±10% on the parameter defined as “Raman scattering per micrometer”. This tolerance value is compatible with a misalignment between the chip and the probe of ±400µm. A tilt of ±1.5° between the chip and the probe can also be acceptable. The Raman probe can be used with an excitation wavelength different from 785 nm because the variation of this parameter does not affect the performance of the instrument. In chapter 4, I have reported the characterization results of the optics ordered. As proof of concept, I have built the mechanical setup of the Raman probe and I have finally demonstrated its functionality by measuring ethanol and methanol spectra. The optical power over the detection area is 90 mW and experimental spectral results obtained for these two fluids are in agreement with data reported in the literature.
Ho deciso di dedicare la mia attività di ricerca per la tesi di laurea al campo della biofotonica siccome offre la possibilità di migliorare la vita delle persone attraverso lo sviluppo di nuovi dispositivi o applicazioni laser nel campo biomedico. Un esempio sono i "lab-on-chip", una nuova classe di chip che consentono di svolgere differenti analisi biochimiche, su un'area molto piccola. In accordo con statistiche mediche, il cancro è ad oggi la seconda causa di morte nel mondo: questa osservazione è stata una forte motivazione per me, a tal punto da dedicare il mio lavoro di tesi su questo tema. I moderni setup ottici usati per il readout dei lab-on-chip sono spesso ingombranti e limitano la diffusione su larga scala di questi innovativi chip, la loro integrazione potrebbe portare ad aumentarne la diffusione e l’utilizzo, soprattutto in campo medico. Lo scopo di questa tesi di laurea è la progettazione di una sonda per spettrometria Raman basata su fibra ottica, destinata al riconoscimento delle cellule tumorali nei liquidi biologici umani. Tra i diversi metodi di rilevamento ottico, abbiamo deciso di fare affidamento alla spettroscopia Raman essendo una tecnica label-free, infatti i diversi composti chimici vengono identificati rispetto alla loro “impronta Raman”. La sonda Raman proposta in questa tesi è stata progettata per il campo medico in quanto il nostro obiettivo è quello di indagare la presenza di cellule tumorali in bio-fluidi umani come l'urina, il sangue, le lacrime, il sudore e la saliva. Ho focalizzato la mia attenzione sui valori estraibili dalle analisi del sangue e delle urine, indagato il loro legame con le alterazioni delle cellule tumorali e introdotto il concetto di lab-on-chip. Nel capitolo 2 ho illustrato i perché la spettroscopia Raman è uno dei migliori metodo di rilevazione ottica per lab-on-chip partendo dai principi dell'effetto Raman. Ho descritto in dettaglio il Raman lab-on-chip utilizzato in laboratorio parlando anche del suo processo di fabbricazione. Nel capitolo 3 ho introdotto il progetto della sonda Raman basata su fibre ottiche, partendo dal concetto di sonda ottica. Ho discusso il design scelto, cioè il "coaxial probe design”, configurazione che utilizza una fibra ottica single-mode per guidare il segnale di eccitazione sopra l'area di rilevamento del chip e una fibra multimodale per raccogliere il Raman scattering generato. Uno specchio dicroico separa il segnale di eccitazione dal Raman scattering raccolto mentre lenti, specchi e filtri ottici presenti nel progetto sono utilizzati per aumentare il throughput del sistema e il signal-to-noise ratio. Per simulare l’andamento della luce nel mezzo, mi sono affidato ad ASAP (Advanced Systems Analysis Program) per ottenere il layout ottimale, mentre Matlab è stato usato per simulare il Raman scattering nell'area di rilevamento. I risultati ottenuti sono stati importanti per definire la geometria della sonda ottimale che ha una lunghezza di 138,9 mm e una larghezza di 13,3 mm, mentre la distanza di lavoro è di 5 mm. Ho fissato la soglia di tolleranza al ± 10% rispetto al parametro definito "Raman scattering per micrometer". Il disallineamento tollerato tra il chip e la sonda ammonta a ±400μm mentre un tilt accettabile è di ± 1,5 °. La sonda Raman può essere utilizzata con una lunghezza d'onda di eccitazione differente da 785 nm poiché la variazione di questo parametro non influenza le prestazioni dello strumento. Nel capitolo 4 ho riportato i risultati derivanti dalla caratterizzazione delle ottiche ordinate per il progetto. Come proof-of-concept, ho construito il mechanical setup della sonda e dimostrato la funzionalità della sonda Raman misurando con successo lo spettro dell’etanolo e del metanolo. La potenza sopra l'area di rilevamento usata in questi esperimenti è stata di 90 mW e i risultati ottenuti per questi due fluidi sono in accordo con i dati riportati in letteratura.
Development of a photonics probe for on-chip confocal Raman spectroscopy.
BARBIERI, GIANCARLO
2015/2016
Abstract
My research activity for the master thesis has been related to biophotonics looking for a way to help people by developing innovative devices and by improving laser applications in the biomedical field. This area of photonic applications has led to the concept of “lab-on-chip”, a new class of chips that integrate different biochemical analyses that can then be processed on a very small area. According to statistical results, cancer is now the second cause of death in the world: this observation was a strong motivation for focusing my thesis work on this issue. External optical setups for lab-on-chip readout are often bulky and limit a wider use of these innovative devices. Integration of the whole readout equipment has to be pursued for increasing their exploitation in the medical field. Lab-on-chips are usually low-cost, user-friendly and in order to exploit these features it would be important to pave the way to a new portable system for cancer diagnosis. To achieve such goal, we need a laser source, a spectrometer, a suitable lab-on-chip and a probe for optical readout. The aim of this master thesis is the design of a fiberoptic-based Raman probe for cancer cells recognition in human bio-fluids. Among different optical detection methods, we decided to rely on Raman scattering being a label-free technique since chemical compounds can be identified by their unique Raman fingerprint. The investigated Raman probe is designed for the medical field since our aim was to investigate the presence of cancer cells in human bio-fluids such as urine, blood, tears, sweat and saliva. In chapter 1, I have introduced cancer as the disease of the century. I have focused my attention on blood and urine analytes and their link with cancer cells. I have introduced the concept of lab-on-chip, discussing the state-of-art of these chips and the materials used in the manufactural processes. In chapter 2, I have illustrated the reasons why Raman spectroscopy is one of the best optical detection method for lab-on-chip starting from the principles of the Raman effect. I have described own freeform reflector Raman lab-on-chip used in the laboratory and its manufacturing process. In chapter 3, I have discussed the design of a fiberoptic-based Raman probe, starting from the concept of optical probe. I have then shown the so called “coaxial probe design”. This layout uses a single mode optical fiber to guide the excitation signal over the detection area of the chip and one multimode fiber to collect the excited Raman scattering. A dichroic mirror separates the excitation light from the detected signal while lens, mirror and optical filters are used to enhance system throughput and signal-to-noise ratio. For the ray-tracing simulations, I relied on ASAP (Advanced Systems Analysis Program) to get the optimum layout and Matlab for the Raman scattering simulation in the detection area. The numerical results have been important to define the geometry of the optimum probe, which has a length of 138.9 mm and a width of 13.3 mm while the best working distance is 5 mm. I have imposed to ensure a tolerance of ±10% on the parameter defined as “Raman scattering per micrometer”. This tolerance value is compatible with a misalignment between the chip and the probe of ±400µm. A tilt of ±1.5° between the chip and the probe can also be acceptable. The Raman probe can be used with an excitation wavelength different from 785 nm because the variation of this parameter does not affect the performance of the instrument. In chapter 4, I have reported the characterization results of the optics ordered. As proof of concept, I have built the mechanical setup of the Raman probe and I have finally demonstrated its functionality by measuring ethanol and methanol spectra. The optical power over the detection area is 90 mW and experimental spectral results obtained for these two fluids are in agreement with data reported in the literature.È consentito all'utente scaricare e condividere i documenti disponibili a testo pieno in UNITESI UNIPV nel rispetto della licenza Creative Commons del tipo CC BY NC ND.
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https://hdl.handle.net/20.500.14239/20755