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High-throughput spheroid screens using volume, resazurin reduction and acid phosphatase activity

Ivanov, Delyan Pavlov; Grabowska, Anna M.; Garnett, Martin C.

Authors

Delyan Pavlov Ivanov delyan.ivanov@nottingham.ac.uk

ANNA GRABOWSKA anna.grabowska@nottingham.ac.uk
Professor of Cancer Microenvironment

Martin C. Garnett m.garnett@nottingham.ac.uk



Contributors

Daniel F. Gilbert
Editor

Oliver Friedrich
Editor

Abstract

Mainstream adoption of physiologically-relevant three-dimensional models has been slow in the last 50 years due to long, manual protocols with poor reproducibility, high price and closed commercial platforms. This chapter describes high-throughput, low-cost, open methods for spheroid viability assessment which use readily-available reagents and open-source software to analyse spheroid volume, metabolism and enzymatic activity. We provide two ImageJ macros for automated spheroid size determination - for both single images and for images in stacks. We also share an Excel template spreadsheet allowing users to rapidly process spheroid size data, analyse plate uniformity (such as edge effects and systematic seeding errors), detect outliers and calculate dose-response. The methods would be useful to researchers in preclinical and translational research planning to move away from simplistic monolayer studies and explore 3D spheroid screens for drug safety and efficacy without substantial investment in money or time.

Peer Reviewed Peer Reviewed
Volume 1601
Series Title Methods in Molecular Biology
Book Title Cell Viability Assays
ISBN 978-1-4939-6960-9
APA6 Citation Ivanov, D. P., Grabowska, A. M., & Garnett, M. C. (in press). High-throughput spheroid screens using volume, resazurin reduction and acid phosphatase activity. In D. F. Gilbert, & O. Friedrich (Eds.), Cell Viability Assays. Humana Press (Springer). https://doi.org/10.1007/978-1-4939-6960-9_4
DOI https://doi.org/10.1007/978-1-4939-6960-9_4
Keywords alamar blue, viability assays, overlay culture, hanging drop, FiJi ImageJ, image analysis, three-dimensional cell culture, in vitro model, preclinical screening, drug sensitivity
Publisher URL https://link.springer.com/protocol/10.1007%2F978-1-4939-6960-9_4
Copyright Statement Copyright information regarding this work can be found at the following address: http://creativecommons.org/licenses/by/4.0
Additional Information The publisher has agreed open access publication via email:


From: Suchy, Monica, Springer US
Sent: 22 May 2017 16:58
To: Ivanov Delyan; AgnesFelema, Anbazhagan
Subject: RE: Springer BWF Book_Gilbert_385089_1_En_Cell Viability Assays//Introductory mail

Dear Delyan,

The book already published so I am going to have to check with Comp Copies and see if the order was processed yet. If it was I will request that they send out a new copy to your home address.

I was unaware of the previous conversation, I spoke with Patrick and in this instance, you have our permission.

Best,
Monica

From: Delyan Ivanov [mailto:Delyan.Ivanov@nottingham.ac.uk]
Sent: Monday, May 22, 2017 3:13 AM
To: Suchy, Monica, Springer US; AgnesFelema, Anbazhagan
Subject: Re: Springer BWF Book_Gilbert_385089_1_En_Cell Viability Assays//Introductory mail


Hi Monica,

Thank you for the reply. I have moved house in the mean time - can the book go to my new address?

Delyan Ivanov
3A Chapel Street
Buxton, SK176HX
UK

In my communication with Oliver and Daniel, they contacted Patrick Marton from Springer US and he mentioned that Springer may be willing to make an exemption for my chapter and allow me to self-archive earlier than the 12 month embargo as per the email below.

Would you or one of your colleagues be able to allow early self-archiving for my chapter?

Best wishes,
Delyan


From: Oliver Friedrich
Sent: 17 July 2016 14:16
To: Delyan Ivanov; Dr. Daniel Gilbert
Subject: Re: Methods in Molecular biology book chapter

Dear Delyan,

thank you very much for submitting your manuscript. Daniel and I will be in due course of editing the chapters within the next four weeks and coe back to everyone for editorial feedback if changes are required.
Regarding your previous questions, I was in contact with Patrick Marton from Springer US and this is what he came back with:
For the first question about the supplementary files -- we can actually accept supplementary files -- we publish them on SpringerLink along with the book. The files are linked from the abstract page for the chapter. What I would like to do if the author is amendable is to have him do as he suggests (publish them on an open database with a link out) while also having him submit them with the chapter so we can publish them online. Doing it both ways provides all readers (electronic and print) easy access to the files.


The second question is a bit trickier as we don't have an open access option for single book chapters, only full volumes (which is cost prohibitive). We do have a self archiving policy in place for authors who are required to submit their chapters to repositories (such as PubMedCentral). The policy calls for a one year embargo on submitting the final author version of the chapter, but if that's too long for this author, I'd be happy to make an exception and grant permission to archive it post publication if that works?

I think this looks very accommodating and I suggest, we finalize thos issue on the 'exception' once we have the chapter in its final form, ok?

Cheers,

Oliver


From: Suchy, Monica, Springer US
Sent: 16 May 2017 14:01
To: Ivanov Delyan; AgnesFelema, Anbazhagan
Subject: RE: Springer BWF Book_Gilbert_385089_1_En_Cell Viability Assays//Introductory mail

Dear Dr. Ivanov,

Here is out policy for self-archiving: http://www.springer.com...-archiving-policy/2124.

Per the agreements the authors signed, each corresponding author will receive a copy of the book. These generally arrive about 5-6 weeks after the book is published. All of the chapter authors are entitled to a 40% discount on everything Springer publishes. In order to get this, they need to go to the follow url and fill out the information:http://www.springer.com...ID=0-154102-12-99823-0.

Best,
Monica

Files

AAM Ivanov_D_Methods in Mol Biology Vol Res APH 3D_Springer.pdf (969 Kb)
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Copyright Statement
Copyright information regarding this work can be found at the following address: http://creativecommons.org/licenses/by/4.0





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