aid classic elispot plate reader  (Cellular Technology Ltd)

Journal: iScience

Article Title: Extensive evolution and T cell escape by SARS-CoV-2 in a 2.5-year persistent infection of an immunocompromised host

doi: 10.1016/j.isci.2026.114917

Figure Lengend Snippet: Magnitude and specificity of T cell memory responses generated by 899-day infection (A) Ex vivo IFNγ-ELISpot well images. DMSO control unstimulated wells and wells stimulated with overlapping peptides covering S1 and S2 region of Spike (duplicates shown), or single peptides S51 and S134. (B) Total magnitude of SARS-CoV-2-specific memory T cell response to structural proteins Spike, membrane (M), nucleoprotein (NP) and open reading frame 3a (ORF3a) and RTC proteins (NSP7, NSP12 polymerase, and NSP13 helicase) colored by protein targeted and measured after persistent infection (>900 days). (C and D) Total magnitude of SARS-CoV-2-specific T cell response (C) or T cell response to Structural and RTC proteins (D) in pre-pandemic samples (pre-August 2019), in exposed healthcare workers (HCW) who remained seronegative, including abortive infections, and in HCW with laboratory confirmed SARS-CoV-2 infections (samples 4 months post-exposure/infection in June to July 2020) for comparison to T cell response in persistently infected patient. (E) Ratio of the magnitude of the T cell response to RTC/structural T cells. Percentage of cohort with a response above 1 (stronger response to RTC than structural proteins) shown below. (F) Magnitude of T cell response to a pool of epitopes from Flu, EBV, and CMV. (B–E) Subset of data previously published in Swadling et al. (A–F) IFNγ-ELISpot. (C and D) Box and Whisker, Tukey. (C–F) Statistical analysis was performed using Kruskal-Wallis tests with Dunn’s correction. ∗ p ≤ 0.05; ∗∗ p ≤ 0.01; ∗∗∗ p ≤ 0.001; ∗∗∗∗ p < 0.0001. (E and F) Bars, geomean.

Article Snippet: Plates were washed in double-distilled H 2 O and left to dry overnight before being read on the AID classic ELISpot plate reader (Immunospot S6 Universal M2 ELISpot Reader).

Techniques: Generated, Infection, Ex Vivo, Enzyme-linked Immunospot, Control, Membrane, Comparison, Whisker Assay

Journal: iScience

Article Title: Extensive evolution and T cell escape by SARS-CoV-2 in a 2.5-year persistent infection of an immunocompromised host

doi: 10.1016/j.isci.2026.114917

Figure Lengend Snippet: Failure of host T cells to recognize emerging virus (A) Ex vivo magnitude of the T cell response to individual ancestral sequence peptides in which mutation arose over persistent infection by IFNγ-ELISpot. (B) Magnitude of the CD4 and CD8 T cell responses after 10-day in vitro peptide expansion with individual ancestral sequence peptides. Percentage of CD4 and CD8 producing IFNγ, TNF, or both are shown. (C) Magnitude of IFNγ+, IFNγ+TNF+, and CTV lo IFNγ+ CD4 and CD8 T cells after 8-day expansion with a pool of 12 peptides corresponding to ancestral sequence epitopes or variant sequence epitopes containing mutations in the virus isolated at day 899 of infection. (D) Magnitude of the IFNγ+TNF+ CD4 or CD8 T cell response after 10-day in vitro peptide stimulation with individual epitopes using ancestral sequence or variant sequence peptides for culture and re-stimulation on day 9. Summary data showing all T cell responses that were detectable (left) or shown for individual peptides (right) for CD4 then CD8 T cells. Duplicate and triplicate stimulations are shown for CD8 T cells for M14 and NSP2, respectively. (D) Bars, mean. Statistical analysis was performed using Kruskal-Wallis tests with Dunn’s correction. ∗ p ≤ 0.05; ∗∗∗ p ≤ 0.001. Peptide sequences shown in .

Article Snippet: Plates were washed in double-distilled H 2 O and left to dry overnight before being read on the AID classic ELISpot plate reader (Immunospot S6 Universal M2 ELISpot Reader).

Techniques: Virus, Ex Vivo, Sequencing, Mutagenesis, Infection, Enzyme-linked Immunospot, In Vitro, Variant Assay, Isolation

Journal: bioRxiv

Article Title: ULK1 drives NDP52-mediated selective autophagic degradation of MHC-I to promote immune evasion in HPV-positive head and neck cancer

doi: 10.64898/2026.03.14.711071

Figure Lengend Snippet: Immune cell profiling of tumor tissues from C57BL/6 mice injected with mEERL/sg-Scr or mEERL/sg2- Ulk1 (n=4/group). Isolated single cell suspensions were stained with an antibody cocktail and analyzed by flow cytometry. Percentages of total T cells of CD45 + (A), CD8 + T (B), and CD4 + T cells (C) of total CD3 + cells are shown. T cell profiling of tumor draining lymph nodes (TDLNs) from mice injected with either mEERL/sg-Scr or mEERL/sg- Ulk1 was isolated 21 days post-injection (n=3/group). Single-cell suspensions were stained with an antibody cocktail containing Zombie NIR, anti-CD45, anti-CD3, anti-CD4, anti-CD8, anti-CD44, and anti-CD62L antibodies. Representative bar graphs showing percentages of total T cells (D), CD4 + T (E), and CD8 + T cells (F), effector memory CD8 + T cells (G), and naïve CD8 + T cells (H). P -value was determined by One-way ANOVA. * p <0.05, ** p <0.01, *** p <0.001, **** p <0.0001. Cytolytic activity of isolated CD8 + T cells co-cultured with mEERL cells at a 50:1, 10:1, or 2:1 T cell: tumor cell ratios were measured by LDH assay (I). Effector functions of isolated CD8 + T cells were evaluated via IFNγ production measured by ELISA (J) and ELISpot (K and L). The number of spot-forming cells was counted using NIH ImageJ. All data shown are mean ± SEM and representative of at least 3 biological replicates. P -value was determined using Student’s t- Test. *p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001.

Article Snippet: IFN-γ production was determined using the Mouse IFN-γ ELISpot Development Module (R&D Systems, No. SEL485), visualized using the ELISpot Blue Color Module (Strep-AP and BCIP-BNBT) (R&D Systems, No. SEL002), and imaged using the ImmunoSpot Analyzer (CTL).

Techniques: Injection, Isolation, Single Cell, Staining, Flow Cytometry, Activity Assay, Cell Culture, Lactate Dehydrogenase Assay, Enzyme-linked Immunosorbent Assay, Enzyme-linked Immunospot