Rapid counting and spectral sorting of live coral larvae using large-particle flow cytometry

Abstract Research with coral embryos and larvae often requires laborious manual counting and sorting of individual specimens, usually via microscopy. Because many coral species spawn only once per year during a narrow temporal window, sample processing is a time-limiting step for research on the ear...

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Autores principales:	Carly J. Randall, Justin E. Speaks, Claire Lager, Mary Hagedorn, Lyndon Llewellyn, Rock Pulak, Julia Thompson, Line K. Bay, David Mead, Andrew J. Heyward, Andrew P. Negri
Formato:	article
Lenguaje:	EN
Publicado:	Nature Portfolio 2020
Materias:	Medicine R Science Q
Acceso en línea:	https://doaj.org/article/9adccfefcb734ed083ce67846a2488b7
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spelling	oai:doaj.org-article:9adccfefcb734ed083ce67846a2488b72021-12-02T16:06:40ZRapid counting and spectral sorting of live coral larvae using large-particle flow cytometry10.1038/s41598-020-69491-02045-2322https://doaj.org/article/9adccfefcb734ed083ce67846a2488b72020-07-01T00:00:00Zhttps://doi.org/10.1038/s41598-020-69491-0https://doaj.org/toc/2045-2322Abstract Research with coral embryos and larvae often requires laborious manual counting and sorting of individual specimens, usually via microscopy. Because many coral species spawn only once per year during a narrow temporal window, sample processing is a time-limiting step for research on the early life-history stages of corals. Flow cytometry, an automated technique for measuring and sorting particles, cells, and cell-clusters, is a potential solution to this bottleneck. Yet most flow cytometers do not accommodate live organisms of the size of most coral embryos (> 250 µm), and sample processing is often destructive. Here we tested the ability of a large-particle flow cytometer with a gentle pneumatic sorting mechanism to process and spectrally sort live and preserved Montipora capitata coral embryos and larvae. Average survival rates of mechanically-sorted larvae were over 90% and were comparable to those achieved by careful hand-sorting. Preserved eggs and embryos remained intact throughout the sorting process and were successfully sorted based on real-time size and fluorescence detection. In-line bright-field microscopy images were captured for each sample object as it passed through the flow-cell, enabling the identification of early-stage embryos (2-cell to morula stage). Samples were counted and sorted at an average rate of 4 s larva−1 and as high as 0.2 s larva−1 for high-density samples. Results presented here suggest that large-particle flow cytometry has the potential to significantly increase efficiency and accuracy of data collection and sample processing during time-limited coral spawning events, facilitating larger-scale and higher-replication studies with an expanded number of species.Carly J. RandallJustin E. SpeaksClaire LagerMary HagedornLyndon LlewellynRock PulakJulia ThompsonLine K. BayDavid MeadAndrew J. HeywardAndrew P. NegriNature PortfolioarticleMedicineRScienceQENScientific Reports, Vol 10, Iss 1, Pp 1-11 (2020)
institution	DOAJ
collection	DOAJ
language	EN
topic	Medicine R Science Q
spellingShingle	Medicine R Science Q Carly J. Randall Justin E. Speaks Claire Lager Mary Hagedorn Lyndon Llewellyn Rock Pulak Julia Thompson Line K. Bay David Mead Andrew J. Heyward Andrew P. Negri Rapid counting and spectral sorting of live coral larvae using large-particle flow cytometry
description	Abstract Research with coral embryos and larvae often requires laborious manual counting and sorting of individual specimens, usually via microscopy. Because many coral species spawn only once per year during a narrow temporal window, sample processing is a time-limiting step for research on the early life-history stages of corals. Flow cytometry, an automated technique for measuring and sorting particles, cells, and cell-clusters, is a potential solution to this bottleneck. Yet most flow cytometers do not accommodate live organisms of the size of most coral embryos (> 250 µm), and sample processing is often destructive. Here we tested the ability of a large-particle flow cytometer with a gentle pneumatic sorting mechanism to process and spectrally sort live and preserved Montipora capitata coral embryos and larvae. Average survival rates of mechanically-sorted larvae were over 90% and were comparable to those achieved by careful hand-sorting. Preserved eggs and embryos remained intact throughout the sorting process and were successfully sorted based on real-time size and fluorescence detection. In-line bright-field microscopy images were captured for each sample object as it passed through the flow-cell, enabling the identification of early-stage embryos (2-cell to morula stage). Samples were counted and sorted at an average rate of 4 s larva−1 and as high as 0.2 s larva−1 for high-density samples. Results presented here suggest that large-particle flow cytometry has the potential to significantly increase efficiency and accuracy of data collection and sample processing during time-limited coral spawning events, facilitating larger-scale and higher-replication studies with an expanded number of species.
format	article
author	Carly J. Randall Justin E. Speaks Claire Lager Mary Hagedorn Lyndon Llewellyn Rock Pulak Julia Thompson Line K. Bay David Mead Andrew J. Heyward Andrew P. Negri
author_facet	Carly J. Randall Justin E. Speaks Claire Lager Mary Hagedorn Lyndon Llewellyn Rock Pulak Julia Thompson Line K. Bay David Mead Andrew J. Heyward Andrew P. Negri
author_sort	Carly J. Randall
title	Rapid counting and spectral sorting of live coral larvae using large-particle flow cytometry
title_short	Rapid counting and spectral sorting of live coral larvae using large-particle flow cytometry
title_full	Rapid counting and spectral sorting of live coral larvae using large-particle flow cytometry
title_fullStr	Rapid counting and spectral sorting of live coral larvae using large-particle flow cytometry
title_full_unstemmed	Rapid counting and spectral sorting of live coral larvae using large-particle flow cytometry
title_sort	rapid counting and spectral sorting of live coral larvae using large-particle flow cytometry
publisher	Nature Portfolio
publishDate	2020
url	https://doaj.org/article/9adccfefcb734ed083ce67846a2488b7
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Rapid counting and spectral sorting of live coral larvae using large-particle flow cytometry

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