Light Microscopy and Raman Imaging of Carotenoids in Plant Cells In Situ and in Released Carotene Crystals

Tomasz Oleszkiewicz , Marta.Z. Pacia , Ewa Grzebelus , Rafał Barański

Abstract

Light microscopy with a bright field mode offers an easy and fast examination of plant specimen for carotenoid presence in its cells. Using basic techniques such as hand sectioned or squashed preparations, carotenoid-rich chromoplasts can be identified without applying any staining procedure and their localization within the cell, their shape and number can be assessed. More detailed information can be obtained by using Raman spectroscopy which is suitable for the analysis of carotenoids due to their unique Raman spectra and allows semiquantification of their contents. Raman imaging (mapping) can be additionally used to show the distribution of carotenoids within the sample. Raman spectra can be taken from extracted carotenoids but can be also obtained directly from plant tissues or cells as Raman measurements are nondestructive for the sample. Here we describe preparations of intact tissue samples, monolayer cell samples, isolated protoplasts as well as carotene crystals released from chromoplasts that are suitable for subsequent observations using light microscopy and for analysis using Raman spectroscopy.
Author Tomasz Oleszkiewicz (FoBaH / IoPBaB)
Tomasz Oleszkiewicz,,
- Institute of Plant Biology and Biotechnology
, Marta.Z. Pacia
Marta.Z. Pacia,,
-
, Ewa Grzebelus (FoBaH / IoPBaB)
Ewa Grzebelus,,
- Institute of Plant Biology and Biotechnology
, Rafał Barański (FoBaH / IoPBaB)
Rafał Barański,,
- Institute of Plant Biology and Biotechnology
Pages245-260
Publication size in sheets0.75
Book Rodriguez-Concepciónn Manuel, Welsch Ralf (eds.): Plant and Food Carotenoids, Methods and Protocols , Methods in Molecular Biology, 2020, Springer, ISBN 978-1-4939-9951-4, [978-1-4939-9952-1], 421 p., DOI:10.1007/978-1-4939-9952-1
Keywords in EnglishCarrot, Chromoplasts, Bright-field microscopy, Nondestructive measurement, Protoplast, Raman mapping, Spectroscopy
URL https://link.springer.com/protocol/10.1007/978-1-4939-9952-1_19
Languageen angielski
Score (nominal)20
Score sourcepublisherList
Citation count*
Additional fields
FinansowanieThe protocol is partially the result of the research project supported by the National Science Centre in Poland (UMO-2013/09/B/NZ9/02379)
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