Citation
- Authors: A Sokal. et al.
- Year: 2023
- Journal: J Exp Med 220 e20220258
- Applications: in vitro / DNA / FectoPRO
- Cell type: Expi293F
Description: Human embryonic kidney Fibroblast
Known as: Expi 293-F, Expi, HEK-293 Expi
Method
The plasmids coding for recombinant proteins were transiently transfected in Expi293F cells (Thermo Fisher Scientific) using FectoPRO DNA transfection reagent (Polyplus) according to the manufacturer’s instructions. The cells were incubated at 37°C for 5 d and then the culture was centrifuged and the supernatant was concentrated. The proteins were purified from the supernatant by affinity chromatography using His-Trap Excel columns (Cytiva). A final step of size-exclusion chromatography in PBS was also performed using a Superdex200 10/300 (Cytiva).
Abstract
Inborn and acquired deficits of type I interferon (IFN) immunity predispose to life-threatening COVID-19 pneumonia. We longitudinally profiled the B cell response to mRNA vaccination in SARS-CoV-2 naive patients with inherited TLR7, IRF7, or IFNAR1 deficiency, as well as young patients with autoantibodies neutralizing type I IFNs due to autoimmune polyendocrine syndrome type-1 (APS-1) and older individuals with age-associated autoantibodies to type I IFNs. The receptor-binding domain spike protein (RBD)-specific memory B cell response in all patients was quantitatively and qualitatively similar to healthy donors. Sustained germinal center responses led to accumulation of somatic hypermutations in immunoglobulin heavy chain genes. The amplitude and duration of, and viral neutralization by, RBD-specific IgG serological response were also largely unaffected by TLR7, IRF7, or IFNAR1 deficiencies up to 7 mo after vaccination in all patients. These results suggest that induction of type I IFN is not required for efficient generation of a humoral response against SARS-CoV-2 by mRNA vaccines.
