2021
Multimodal Imaging of 3D Cell Aggregates
PREISLER, Jan, Markéta MACHÁLKOVÁ, Barbora ADAMOVÁ, Jarmila NAVRÁTILOVÁ, Marek STIBOREK et. al.Základní údaje
Originální název
Multimodal Imaging of 3D Cell Aggregates
Autoři
PREISLER, Jan, Markéta MACHÁLKOVÁ, Barbora ADAMOVÁ, Jarmila NAVRÁTILOVÁ, Marek STIBOREK, Stanislava MELIORISOVÁ, Jiří KROUPA, Pavel HOUŠKA, Jan MICHÁLEK, Karel ŠTĚPKA, Katarzyna Anna RADASZKIEWICZ, Adam PRUŠKA, Viktor KANICKÝ, Michal KOZUBEK a Jan ŠMARDA
Vydání
MSB 2021, 2021
Další údaje
Jazyk
čeština
Typ výsledku
Vyžádané přednášky
Obor
10406 Analytical chemistry
Stát vydavatele
Česká republika
Utajení
není předmětem státního či obchodního tajemství
Organizační jednotka
Přírodovědecká fakulta
Klíčová slova anglicky
multimodal, imaging, mass spectrometry, 3D cell aggregates
Změněno: 4. 1. 2022 12:35, prof. Mgr. Jan Preisler, Ph.D.
V originále
Spheroids, 3D cell aggregates are popular for studies of drug uptake and diffusion. Proliferating and apoptotic cells in the spheroids can be visualized using immunofluorescence microscopy (IFM). Distribution of drugs, which often do not exhibit specific fluorescence, has to be revealed using another technique, such as matrix-assisted laser desorption/ionization mass spectrometry imaging (MALDI MSI). To evaluate the efficacy of a potential anti-cancer drug perifosine within spheroids of colorectal carcinoma, images of drug abundance from MALDI MSI, and fluorescently visualized proliferating and/or apoptotic cells must be precisely colocalized and carefully compared. Approximately 1-mm spheroids (HT-29 cell line) treated with perifosine were imaged with MALDI MS, the matrix was washed away and cell viability was inspected with IFM. Alternatively, laser ablation coupled to inductively coupled plasma mass spectrometry (LA - ICP MS) was applied for more sensitive viability visualization using immunostaining with 20-nm gold nanoparticles as the tags. The overlay of MALDI MS and IFM spheroid images required fiducial-based coregistration because of the lack of any morphological features in the spheroids. The spatial relationship between the MALDI MS and IFM intensities was evaluated based on the respective intensities along equidistant layers, or “peels” of the entire spheroid, starting from the spheroid boundary. The accurate colocalization of MALDI MS and IFM maps and subsequent peeling data analysis showed a limited penetration of perifosine into spheroids within 24 hours and allowed differentiation between apoptosis resulting from hypoxia/nutrient deprivation and drug exposure. As expected, viability assays from IFM and LA - ICP MS for markers of cell proliferation and apoptosis revealed a higher signal of the apoptotic marker in the spheroid core and a higher signal of proliferative cells on the spheroid edge. We thank to CSF (21-12262S) and GAMU (MUNI/G/0974/2016).
Anglicky
Spheroids, 3D cell aggregates are popular for studies of drug uptake and diffusion. Proliferating and apoptotic cells in the spheroids can be visualized using immunofluorescence microscopy (IFM). Distribution of drugs, which often do not exhibit specific fluorescence, has to be revealed using another technique, such as matrix-assisted laser desorption/ionization mass spectrometry imaging (MALDI MSI). To evaluate the efficacy of a potential anti-cancer drug perifosine within spheroids of colorectal carcinoma, images of drug abundance from MALDI MSI, and fluorescently visualized proliferating and/or apoptotic cells must be precisely colocalized and carefully compared. Approximately 1-mm spheroids (HT-29 cell line) treated with perifosine were imaged with MALDI MS, the matrix was washed away and cell viability was inspected with IFM. Alternatively, laser ablation coupled to inductively coupled plasma mass spectrometry (LA - ICP MS) was applied for more sensitive viability visualization using immunostaining with 20-nm gold nanoparticles as the tags. The overlay of MALDI MS and IFM spheroid images required fiducial-based coregistration because of the lack of any morphological features in the spheroids. The spatial relationship between the MALDI MS and IFM intensities was evaluated based on the respective intensities along equidistant layers, or “peels” of the entire spheroid, starting from the spheroid boundary. The accurate colocalization of MALDI MS and IFM maps and subsequent peeling data analysis showed a limited penetration of perifosine into spheroids within 24 hours and allowed differentiation between apoptosis resulting from hypoxia/nutrient deprivation and drug exposure. As expected, viability assays from IFM and LA - ICP MS for markers of cell proliferation and apoptosis revealed a higher signal of the apoptotic marker in the spheroid core and a higher signal of proliferative cells on the spheroid edge. We thank to CSF (21-12262S) and GAMU (MUNI/G/0974/2016).
Návaznosti
| GA21-12262S, projekt VaV |
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| MUNI/G/0974/2016, interní kód MU |
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