Simultaneous analysis of antigen‐specific B and T cells after SARS‐CoV‐2 infection and vaccination

Abstract

Conventional methods for quantifying and phenotyping antigen‐specific lymphocytes can rapidly deplete irreplaceable specimens. This is due to the fact that antigen‐specific T and B cells have historically been analyzed in independent assays each requiring millions of cells. A technique that facilitates the simultaneous detection of antigen‐specific T and B cells would allow for more thorough immune profiling with significantly reduced sample requirements. To this end, we developed the B and T cell tandem lymphocyte evaluation (BATTLE) assay, which allows for the simultaneous identification of SARS‐CoV‐2 Spike reactive T and B cells using an activation induced marker (AIM) T cell assay and dual‐color B cell antigen probes. Using this assay, we demonstrate that antigen‐specific B and T cell subsets can be identified simultaneously using conventional flow cytometry platforms and provide insight into the differential effects of mRNA vaccination on B and T cell populations following natural SARS‐CoV‐2 infection.

Activation‐induced marker assays for identification of Trypanosoma cruzi‐specific CD4 or CD8 T cells in chronic Chagas disease patients

“…Next, we evaluated the co‐upregulation of the 10 AIM combinations described above in CD8 + T‐cell population from PBMC of CCD and NI individuals after T . cruzi stimulation at both time points, 12 and 24 h. By mirroring our approach used for CD4, incubation times were selected on the basis of the activation markers kinetic and previous reports on AIM assays [11, 15–18, 20, 21, 25, 35, 40].…”

“…SARS‐CoV‐2‐specific CD4 + T lymphocytes were detected with OX40/CD137 AIM assay at 24 h stimulation [18]. In the same disease, specific CD4 + T‐cell responses were quantified in PBMC from COVID‐19 patients by OX40/CD40L [26], OX40/CD69 [25] or CD69/CD137 [24] AIM combinations after 24 h of stimulation.…”

“…Gate-pathway used to select this cell subset is graphed in Figure S1. CCD patients and NI subjects stimulated in vitro with parasite lysate during 12 or 24 h, considering the kinetics of each activation markers and stimulation time used in previous studies [11,[13][14][15][16][17][18][19][20][21][22][23][24][25][26]35]. Although the expression profile of these molecules is variable upon antigen stimulation, and some of them peak later, that is, OX40 at 48 h [36], these time constraints were chosen to capture their simultaneous upregulation in the AIM combinations.…”

“…In the present study, we decided to expand on our previous result by using other combinations of surface molecular markers to detect T. cruzi-stimulated CD4 + T cells and also to find any that enable us to identify parasite-activated CD8 + T cells. Based on an increase number of reports [11,[14][15][16][17][18][19][20][21][22][23][24][25][26], the markers CD69, CD40L, CD137, PD-L1, CD11a, CD49d, CD38, HLA-DR as well as OX40 and CD25 were included in different flow cytometry panels for a dual AIM assay. For T-cell agnostic activation, PBMC from patients with CCD or non-infected donors were in vitro stimulated for 12 and 24 h with trypomastigote lysates of different strains (Sylvio, CL Brener and Dm28c).…”

Anti-dsDNA B-Cell ELISpot as a Monitoring and Flare Prediction Tool in SLE Patients

“…Additionally, in recent years, some authors had studied the detection of antigen-specific B-cells by flow cytometry [ 36 , 37 , 38 ]. Since flow cytometry techniques are available in most immunology laboratories, it might be of interest to compare the presence of anti-dsDNA secreting cells detected by SLE-ELISpot with dsDNA-specific B-cells detected by flow cytometry.…”