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DTSTART:20220614T130000Z
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TRANSP:OPAQUE
LOCATION:ICFO Auditorium and Online (Teams)
SUMMARY:ICFO | MATTEO BERNARDELLO 
CLASS:PUBLIC
DESCRIPTION:The observation of biological processes in their native environ
 ments is of critical importance for life science. While substantial inform
 ation can be derived from the examination of in-vitro biological samples\,
  in-vivo studies are necessary to reveal the complexity of the dynamics ha
 ppening in real-time within a living organism. Between the possible biolog
 ical model choices\, vertebrates represent an important family due to the 
 various characteristics they share with the human organism. The developmen
 t of an embryo\, the effect of a drug\, the interaction between the immune
  system and pathogens\, and the cellular machinery activities are all exam
 ples of highly-relevant applications requiring in-vivo observations on bro
 adly used vertebrate models such as the zebrafish and the mouse.\nTo perfo
 rm such observations\, appropriate devices have been devised. Fluorescence
  microscopy is one of the main approaches through which specific sample st
 ructures can be detected and registered in high-contrast images. Through m
 icro-injections or transgenic lines\, a living specimen can express fluore
 scence and can be imaged through such microscopes. Various fluorescence mi
 croscopy techniques have been developed\, such as Widefield Microscopy (WM
 ) and Laser Scanning Confocal Microscopy (LSCM). In WM the entire sample i
 s visualized in a single 2D image\, therefore losing the depth information
 \, while LSCM can recover the 3D information of the sample but with inhere
 nt limitations\, such as phototoxicity and limited imaging speed.\nIn the 
 last two decades\, Light-Sheet Fluorescence Microscopy (LSFM) emerged as a
  technique providing fast and 3D imaging\, while minimizing collateral dam
 ages to the specimen. However\, due to the particular configuration of the
  microscope&rsquo\;s components\, LSFM setups are normally optimized for a
  single application. Also\, sample management is not trivial\, as controll
 ing the specimen positioning and keeping it alive for a long time within t
 he microscope needs dedicated environmental conditioning.\nIn this thesis\
 , I aimed at advancing the imaging flexibility of LSFM\, with particular a
 ttention to sample management. The conjugation of these aspects enabled no
 vel observations and applications on living vertebrate samples.&nbsp\;\nIn
  Chapter 1\, a brief review of the concepts employed within this thesis is
  presented\, also pointing to the main challenges that the thesis aims to 
 solve.\nIn Chapter 2\, a new design for multimodal LSFM is presented\, whi
 ch enables performing different experiments with the same instrument. Part
 icularly\, high-throughput studies would benefit from this imaging paradig
 m\, conjugating the need for fast and reproducible mounting of multiple sa
 mples with the opportunity to image them in 3D. Additionally\, from this d
 esign\, a transportable setup has also been implemented.\nWith these syste
 ms\, I studied the dynamics of the yolk&rsquo\;s microtubule network of ze
 brafish embryos\, describing novel features and underlining the importance
  of live imaging to have a whole view of the sample&rsquo\;s peculiarities
 . This is described in Chapter 3.\nFurther applications on challenging liv
 e samples have been implemented\, monitoring the macrophage recruitment in
  zebrafish and the development of mouse embryos. For these applications\, 
 described in Chapter 4\, I devised specific mounting protocols for the sam
 ples\, keeping them alive during the imaging sessions.\nIn Chapter 5\, an 
 additional LSFM system is described\, which allows for recording the sub-c
 ellular machinery in a living vertebrate sample\, while avoiding its damag
 e thanks to the devised sample mounting. Through this\, single-molecule mi
 croscopy (SMM) studies\, normally performed on cultured cells\, can be ext
 ended to the nuclei of living zebrafish embryos\, which better recapitulat
 e the native environment where biological processes take place.\nFinally\,
  Chapter 6 recapitulates the conclusions\, the impacts\, future integratio
 ns\, and experimental procedures that would be enabled by the work resumed
  in this thesis.\n&nbsp\;\nThesis Director: Dr Pablo Loza\nThesis Co-Direc
 tor: Dr Emilio J. Gualda
DTSTAMP:20260407T052605Z
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