Enhancing the sensitivity of assays to detect T cell reactivity: the effect of cell separation and cryopreservation media.

Abstract:

:The identification of sensitive assay formats capable of distinguishing islet autoreactive T cells directly ex vivo in blood is a major goal in type 1 diabetes research. Whereas much effort has been made to identify suitable assay formats, relatively little attention has been paid to optimizing the quality of cell preparations used. To address this issue we investigated the role of two key variables in the preparation of peripheral blood mononuclear cells (PBMCs): the separation media used in density gradient centrifugation and the method of cryopreservation. PBMCs were prepared from a single individual using different protocols and tested for responses to suboptimal concentrations of tetanus toxoid, representing a low frequency recall response. Significant differences were observed in T cell responses dependent upon the selection of the separation media and cryopreservation methods used, indicating that relatively small differences in preparation of PBMCs can have measurable effects on assay sensitivity.

journal_name

Ann N Y Acad Sci

authors

Tree TI,Roep BO,Peakman M

doi

10.1196/annals.1337.005

subject

Has Abstract

pub_date

2004-12-01 00:00:00

pages

26-32

eissn

0077-8923

issn

1749-6632

pii

1037/1/26

journal_volume

1037

pub_type

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