Display Settings:

Format

Send to:

Choose Destination
See comment in PubMed Commons below
Proc Natl Acad Sci U S A. 2012 Mar 6;109(10):3885-90. doi: 10.1073/pnas.1111205109. Epub 2012 Feb 21.

Rapid, efficient functional characterization and recovery of HIV-specific human CD8+ T cells using microengraving.

Author information

  • 1Department of Chemical Engineering, Massachusetts Institute of Technology, Cambridge, MA 02139, USA.

Abstract

The nature of certain clinical samples (tissue biopsies, fluids) or the subjects themselves (pediatric subjects, neonates) often constrain the number of cells available to evaluate the breadth of functional T-cell responses to infections or therapeutic interventions. The methods most commonly used to assess this functional diversity ex vivo and to recover specific cells to expand in vitro usually require more than 10(6) cells. Here we present a process to identify antigen-specific responses efficiently ex vivo from 10(4)-10(5) single cells from blood or mucosal tissues using dense arrays of subnanoliter wells. The approach combines on-chip imaging cytometry with a technique for capturing secreted proteins--called "microengraving"--to enumerate antigen-specific responses by single T cells in a manner comparable to conventional assays such as ELISpot and intracellular cytokine staining. Unlike those assays, however, the individual cells identified can be recovered readily by micromanipulation for further characterization in vitro. Applying this method to assess HIV-specific T-cell responses demonstrates that it is possible to establish clonal CD8(+) T-cell lines that represent the most abundant specificities present in circulation using 100- to 1,000-fold fewer cells than traditional approaches require and without extensive genotypic analysis a priori. This rapid (<24 h), efficient, and inexpensive process should improve the comparative study of human T-cell immunology across ages and anatomic compartments.

PMID:
22355106
[PubMed - indexed for MEDLINE]
PMCID:
PMC3309713
Free PMC Article

Images from this publication.See all images (4)Free text

Fig. 1.
Fig. 2.
Fig. 3.
Fig. 4.
PubMed Commons home

PubMed Commons

0 comments
How to join PubMed Commons

    Supplemental Content

    Icon for HighWire Icon for PubMed Central
    Loading ...
    Write to the Help Desk