Stable phenotype of B‐cell subsets following cryopreservation and thawing of normal human lymphocytes stored in a tissue biobank. (20th October 2014)

• Record Type: Journal Article
• Authors: Rasmussen, Simon Mylius; Bilgrau, Anders Ellern; Schmitz, Alexander; Falgreen, Steffen; Bergkvist, Kim Steve; Tramm, Anette Mai; Bæch, John; Jacobsen, Chris Ladefoged; Gaihede, Michael; Kjeldsen, Malene Krag; Bødker, Julie Støve; Dybkær, Karen; Bøgsted, Martin; Johnsen, Hans Erik
• Is Part Of: Cytometry. Volume 88:Number 1(2015)
• Page Start: 40
• Page End: 49
• Publication Date: 2014-10-20
• Subjects: Flow cytometry -- Diagnostic use -- Periodicals; Cytodiagnosis -- Periodicals; 616.07582
• DOI: 10.1002/cyto.b.21192
• Languages: English
• ISSNs: 1552-4949

Abstract

<abstract abstract-type="main"> <title> <x xml:space="preserve">Abstract</x> </title> <sec id="cytob21192-sec-0001" sec-type="section"> <title>Background</title> <p>Cryopreservation is an acknowledged procedure to store vital cells for future biomarker analyses. Few studies, however, have analyzed the impact of the cryopreservation on phenotyping.</p> </sec> <sec id="cytob21192-sec-0002" sec-type="section"> <title>Methods</title> <p>We have performed a controlled comparison of cryopreserved and fresh cellular aliquots prepared from individual healthy donors. We studied circulating B‐cell subset membrane markers and global gene expression, respectively by multiparametric flow cytometry and microarray data. Extensive statistical analysis of the generated data tested the concept that "overall, there are no phenotypic differences between cryopreserved and fresh B‐cell subsets." Subsequently, we performed an uncontrolled comparison of tonsil tissue samples.</p> </sec> <sec id="cytob21192-sec-0003" sec-type="section"> <title>Results</title> <p>By multiparametric flow analysis, we documented no significant changes following cryopreservation of subset frequencies or membrane intensity for the differentiation markers CD19, CD20, CD22, CD27, CD38, CD45, and CD200. By gene expression profiling following cryopreservation, across all samples, only 16 out of 18708 genes were significantly up or down regulated, including <italic>FOSB, KLF4, RBP7, ANXA1</italic> or <italic>CLC, DEFA3</italic>, respectively. Implementation of cryopreserved tissue in our research program allowed us to present a performance analysis, by comparing cryopreserved and fresh tonsil tissue. As expected, phenotypic differences were identified, but to an extent that did not affect the performance of the cryopreserved tissue to generate specific B‐cell subset associated gene signatures and assign subset phenotypes to independent tissue samples.</p> </sec> <sec id="cytob21192-sec-0004" sec-type="section"> <title>Conclusions</title> <p>We have confirmed our working concept and illustrated the usefulness of vital cryopreserved cell suspensions for phenotypic studies of the normal B‐cell hierarchy; however, storage procedures need to be delineated by tissue‐specific comparative analysis. © 2014 International Clinical Cytometry Society</p> </sec> </abstract>