Spike-specific T-cell Immune Response Assay

ICS staining and flow cytometry were performed as described in detail [12 (link)], except differing in the antigenic target. In brief, PBMCs were incubated with a pool of overlapping 15-mer peptides spanning spike [13 ] at a final concentration of 1 µg/ml of each individual peptide, with brefeldin A (#00-4506-51, eBioscience, San Diego, CA) and monensin (#00-4505-51, eBioscience, San Diego, CA), followed by surface staining CD3-Super Bright 436, CD8-Super Bright 600, CD4 PE-Cy7, and Fixable Aqua viability dye (#62-0037-42, eBioscience, San Diego/CA; #63-0088-42, eBioscience, San Diego/CA; #25-0049-42, San Diego, CA; and #L34957, Invitrogen, Waltham, MA respectively), permeabilization (#00-5523-00, eBioscience, San Diego, CA), and intracellular cytokine staining for interferon (IFN)-ɣ-FITC, IL-2-PerCP-Cy5.5, IL-4-PE, and IL-10-APC (#506504 Biolegend, San Diego, CA; #500322, Biolegend, San Diego, CA; # 130-091-647, Miltenyi Biotec, Bergisch Gladbach, Germany; and #506807, Biolegend, San Diego, CA respectively) for flow cytometric analysis.

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Taus E., Shino M.Y., Ibarrondo F.J., Hausner M.A., Hofmann C, & Yang O.O. (2023). Predominantly defective CD8+ T cell immunity to SARS-CoV-2 mRNA vaccination in lung transplant recipients. Journal of Translational Medicine, 21, 374.

Publication 2023

brefeldin A monensin CD3-Super Bright 436 CD8-Super Bright 600 CD4 PE-Cy7 Fixable Aqua viability dye

Antigenic Brefeldin a Cy5 5 Cytokine Fitc Flow cytometric Il 10 Interferon Intracellular Monensin Peptide

Corresponding Organization :

Other organizations : University of California, Los Angeles

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Variable analysis

independent variables

Pool of overlapping 15-mer peptides spanning spike

dependent variables

Interferon (IFN)-ɣ production
IL-2 production
IL-4 production
IL-10 production

control variables

Brefeldin A
Monensin
CD3-Super Bright 436
CD8-Super Bright 600
CD4 PE-Cy7
Fixable Aqua viability dye

positive controls

Not explicitly mentioned

negative controls

Not explicitly mentioned

Annotations

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