ELISPOTs Produced by CD8 and CD4 Cells Follow Log Normal Size Distribution Permitting Objective Counting

Each positive well in ELISPOT assays contains spots of variable sizes that can range from tens of micrometers up to a millimeter in diameter. Therefore, when it comes to counting these spots the decision on setting the lower and the upper spot size thresholds to discriminate between non-specific bac...

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Main Authors:	Karulin, Alexey Y., Karacsony, Kinga, Zhang, Wenji, Targoni, Oleg S., Moldovan, Ioana, Dittrich, Marcus, Sundararaman, Srividya, Lehmann, Paul V.
Format:	Online
Language:	English
Published:	MDPI 2015
Online Access:	https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4381209/

id	pubmed-4381209
recordtype	oai_dc
spelling	pubmed-43812092015-05-18 ELISPOTs Produced by CD8 and CD4 Cells Follow Log Normal Size Distribution Permitting Objective Counting Karulin, Alexey Y. Karacsony, Kinga Zhang, Wenji Targoni, Oleg S. Moldovan, Ioana Dittrich, Marcus Sundararaman, Srividya Lehmann, Paul V. Article Each positive well in ELISPOT assays contains spots of variable sizes that can range from tens of micrometers up to a millimeter in diameter. Therefore, when it comes to counting these spots the decision on setting the lower and the upper spot size thresholds to discriminate between non-specific background noise, spots produced by individual T cells, and spots formed by T cell clusters is critical. If the spot sizes follow a known statistical distribution, precise predictions on minimal and maximal spot sizes, belonging to a given T cell population, can be made. We studied the size distributional properties of IFN-γ, IL-2, IL-4, IL-5 and IL-17 spots elicited in ELISPOT assays with PBMC from 172 healthy donors, upon stimulation with 32 individual viral peptides representing defined HLA Class I-restricted epitopes for CD8 cells, and with protein antigens of CMV and EBV activating CD4 cells. A total of 334 CD8 and 80 CD4 positive T cell responses were analyzed. In 99.7% of the test cases, spot size distributions followed Log Normal function. These data formally demonstrate that it is possible to establish objective, statistically validated parameters for counting T cell ELISPOTs. MDPI 2015-01-20 /pmc/articles/PMC4381209/ /pubmed/25612115 http://dx.doi.org/10.3390/cells4010056 Text en © 2015 by the authors; licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution license (http://creativecommons.org/licenses/by/4.0/).
repository_type	Open Access Journal
institution_category	Foreign Institution
institution	US National Center for Biotechnology Information
building	NCBI PubMed
collection	Online Access
language	English
format	Online
author	Karulin, Alexey Y. Karacsony, Kinga Zhang, Wenji Targoni, Oleg S. Moldovan, Ioana Dittrich, Marcus Sundararaman, Srividya Lehmann, Paul V.
spellingShingle	Karulin, Alexey Y. Karacsony, Kinga Zhang, Wenji Targoni, Oleg S. Moldovan, Ioana Dittrich, Marcus Sundararaman, Srividya Lehmann, Paul V. ELISPOTs Produced by CD8 and CD4 Cells Follow Log Normal Size Distribution Permitting Objective Counting
author_facet	Karulin, Alexey Y. Karacsony, Kinga Zhang, Wenji Targoni, Oleg S. Moldovan, Ioana Dittrich, Marcus Sundararaman, Srividya Lehmann, Paul V.
author_sort	Karulin, Alexey Y.
title	ELISPOTs Produced by CD8 and CD4 Cells Follow Log Normal Size Distribution Permitting Objective Counting
title_short	ELISPOTs Produced by CD8 and CD4 Cells Follow Log Normal Size Distribution Permitting Objective Counting
title_full	ELISPOTs Produced by CD8 and CD4 Cells Follow Log Normal Size Distribution Permitting Objective Counting
title_fullStr	ELISPOTs Produced by CD8 and CD4 Cells Follow Log Normal Size Distribution Permitting Objective Counting
title_full_unstemmed	ELISPOTs Produced by CD8 and CD4 Cells Follow Log Normal Size Distribution Permitting Objective Counting
title_sort	elispots produced by cd8 and cd4 cells follow log normal size distribution permitting objective counting
description	Each positive well in ELISPOT assays contains spots of variable sizes that can range from tens of micrometers up to a millimeter in diameter. Therefore, when it comes to counting these spots the decision on setting the lower and the upper spot size thresholds to discriminate between non-specific background noise, spots produced by individual T cells, and spots formed by T cell clusters is critical. If the spot sizes follow a known statistical distribution, precise predictions on minimal and maximal spot sizes, belonging to a given T cell population, can be made. We studied the size distributional properties of IFN-γ, IL-2, IL-4, IL-5 and IL-17 spots elicited in ELISPOT assays with PBMC from 172 healthy donors, upon stimulation with 32 individual viral peptides representing defined HLA Class I-restricted epitopes for CD8 cells, and with protein antigens of CMV and EBV activating CD4 cells. A total of 334 CD8 and 80 CD4 positive T cell responses were analyzed. In 99.7% of the test cases, spot size distributions followed Log Normal function. These data formally demonstrate that it is possible to establish objective, statistically validated parameters for counting T cell ELISPOTs.
publisher	MDPI
publishDate	2015
url	https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4381209/
_version_	1613206104829853696

ELISPOTs Produced by CD8 and CD4 Cells Follow Log Normal Size Distribution Permitting Objective Counting

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