|
| Status |
Public on Aug 06, 2024 |
| Title |
D12 T1 Prot_S stimulated |
| Sample type |
RNA |
| |
|
| Source name |
Whole blood - 24 hours - 37°C - anti-CD28+anti-CD49d+SARS-CoV-2 Prot_S
|
| Organism |
Homo sapiens |
| Characteristics |
tissue: Whole blood
date 1st dose bnt162b2: 2021-03-10
date 2nd dose bnt162b2: 2021-03-31
date 3rd dose bnt162b2 (booster): 2021-11-16
date timepoint 0 (pre-booster): 2021-11-16
date timepoint 1 month post-booster: 2021-12-13
date timepoint 3 months post-booster: 2022-02-01
date timepoint 6 months post-booster: 2022-05-04
gender: Female
age at timepoint 0: 35
|
| Treatment protocol |
As published previously (Lauruschkat et al., 2021, J Fungi), 2.7-mL blood collection tubes without anticoagulant were pre-loaded with co-stimulatory antibodies α-CD28 and α-CD49d (10 µg/mL, Miltenyi Biotec, Bergisch Gladbach, Germany) plus either no antigen (negative control) or SARS-CoV-2 spike peptides (Prot_S, 0.6 nmol per peptide/mL, Miltenyi Biotec, Bergisch Gladbach, Germany). 0.5 mL blood lithium-heparin blood was added and stimulation tubes were incubated for 20 – 24 h at 37 °C. Samples were transferred to 1.5 mL tubes and centrifuged at 2000 g for 20 min. After preservation of the supernatant for LegendPlex cytokine assays, the residual cell pellet underwent erythrocyte lysis using Buffer EL (Qiagen, Hilden, Germany) and was then cryopreserved in 1 mL RNAprotect buffer (Qiagen, Hilden, Germany).
|
| Growth protocol |
Not applicable
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA was isolated using the RNeasy Plus Mini Kit according to the manufacturer's instructions (Qiagen, Hilden, Germany).
|
| Label |
n/a
|
| Label protocol |
Hybridisation with labelled probes was performed according to the manufacturer's instructions (Nanostring, Seattle, USA): RNA concentration was normalized to 50 ng in 5 µL water. The lyophilyzed Reporter Code set was reconstitued using 70 µL Hybridisation Buffer. 50 ng of RNA was mixed with 8 µL of Reporter Codeset solution and 2 µL of (ready-to-use) Capture Probe Set.
|
| |
|
| Hybridization protocol |
Hybridisation was performed for 16-24 h at 65 °C.
|
| Scan protocol |
17 µL water was added to the hybridized samples. 30 µL of each sample was loaded onto a Nanostring nCounter Sprint Cartridge and analyzed using an nCounter Sprinter device (Nanostring, Seattle, USA).
|
| Description |
nCounter Immune Exhaustion Panel Reporter CodeSet
|
| Data processing |
Data was analyzed using the nSolver Analysis Software (Nanostring, Seattle, USA), with background thresholding to the median of negative controls and normalization to the panel’s 12 internal reference genes (geometric mean).
mRNA counts normalized to internal reference genes by the nSolver Analysis Software (Nanostring, Seattle, USA).
|
| |
|
| Submission date |
Aug 05, 2024 |
| Last update date |
Aug 06, 2024 |
| Contact name |
Lukas Page |
| Organization name |
University Hospital of Augsburg
|
| Department |
Institute for Laboratory Medicine and Microbiology
|
| Lab |
Reinhard Hoffmann
|
| Street address |
Stenglinstrasse 2
|
| City |
Augsburg |
| State/province |
Bavaria |
| ZIP/Postal code |
86156 |
| Country |
Germany |
| |
|
| Platform ID |
GPL33412 |
| Series (1) |
| GSE273994 |
Antigen-specific T helper cells predict intensity and longevity of immune responses to SARS-CoV-2 booster vaccination |
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