2’3’-cgamp  (InvivoGen)

Journal: iScience

Article Title: cGAS/STING sensing in dendritic cells discriminates between daptomycin sensitive and resistant Staphylococcus aureus clinical isolates

doi: 10.1016/j.isci.2026.115854

Figure Lengend Snippet: DapR MRSA fails to stimulate STING phosphorylation in DC (A) Western blot of cell lysates of MutuDC stimulated with complete media, 10 nmol 3′3′-cGAMP, MOI = 10 of MSSA (D85), DapS (A8819) MRSA, or DapR (A8817) MRSA, for the indicated times. (B) Quantitation of p-STING relative to total STING for stimulations as in “A,” at 90 min post stimulus. Bars show the geometric mean and geometric SD of replicates pooled from three independent experiments ( n = 3). Statistical significance determined using paired t test and reported as indicated by ∗ p ≤ 0.05.

Article Snippet: 3′3′-cGAMP- Cyclic [G(3′,5′)pA(3′,5′)p] , InvivoGen , Cat#: tlrl-nacga.

Techniques: Phospho-proteomics, Western Blot, Quantitation Assay

Journal: iScience

Article Title: cGAS/STING sensing in dendritic cells discriminates between daptomycin sensitive and resistant Staphylococcus aureus clinical isolates

doi: 10.1016/j.isci.2026.115854

Figure Lengend Snippet: MRSA isolates differentially induce 2′3′-cGAMP production in DC and differentially secrete 3′3'-cGAMP in response to stress signals (A) Detection of mammalian 2′3′-cGAMP (pmol/million DC) in MutuDC (lysates) following 1, 2 or 4 h stimulation with DapS (A8819; light blue) MRSA or DapR (A8817; dark blue) MRSA at an MOI of 10. Bars show the mean and SEM of 2′3′-cGAMP detected from three biological replicates ( n = 3). (B and C) Detection of bacterial 3′3′-cGAMP in fresh (B) or freeze-thawed (C) supernatants of DapS (A8819; light blue) MRSA or DapR (A8817; dark blue) MRSA cultured in brain heart infusion (BHI) medium overnight at a density of 4 × 10 8 cfu/ml. (D) Detection of bacterial 3′3′-cGAMP in fresh MutuDC lysates following 2 h of stimulation with DapS (A8819; light blue) MRSA or DapR (A8817; dark blue) at an MOI of 10. Bars show the mean and SEM of 3′3′-cGAMP detected from three biological replicates ( n = 3). (E and F) Bacterial 3′3′-cGAMP production by DapS (A8819; light blue) MRSA or DapR (A8817; dark blue) isolates cultured in BHI culture media (E) or DC culture media (F); with each media alone (solid bars) or supplemented with antibiotics (dashed bars), consisting of penicillin (10 units/ml) and streptomycin (10 μg/mL) and incubated overnight at a density of 8 × 10 6 cfu/ml. Bars indicate 3′3′-cGAMP at 30, 60, 90, and 120 min post incubation, showing the mean and SEM from three biological replicates ( n = 3). (A, B, C, and D) Significance reflective of an unpaired two-tailed t test using Holm Sidak correction, reported as indicated by ∗ p ≤ 0.05 and ∗∗∗ p < 0.0005. (E and F) Paired two-tailed t test where ∗ p < 0.05 and ∗∗ p < 0.005.

Article Snippet: 3′3′-cGAMP- Cyclic [G(3′,5′)pA(3′,5′)p] , InvivoGen , Cat#: tlrl-nacga.

Techniques: Cell Culture, Incubation, Two Tailed Test

Journal: iScience

Article Title: cGAS/STING sensing in dendritic cells discriminates between daptomycin sensitive and resistant Staphylococcus aureus clinical isolates

doi: 10.1016/j.isci.2026.115854

Figure Lengend Snippet: CDN secretion in response to stress by DapS, but not DapR, MRSA In steady state, both DapS and DapR MRSA produce very low levels of 3′3′- CDN. Under stress conditions, bacteria can activate the CBASS defense system, triggering the production of the bacterial CDN 3′3′-cGAMP, a potent mammalian STING agonist. Under stress, DapS MRSA produces plentiful 3′3′-cGAMP, and the weakened bacterial envelope can lead to bacterial DNA release, providing mammalian cGAS stimulus and 2′3′-cGAMP production and further STING activation, which also enhances phagocytosis. The complex cell wall of DapR MRSA provides “innate” protection during stress and prevents the triggering of CBASS to produce CDN.

Article Snippet: 3′3′-cGAMP- Cyclic [G(3′,5′)pA(3′,5′)p] , InvivoGen , Cat#: tlrl-nacga.

Techniques: Bacteria, Activation Assay

Journal: The Journal of Clinical Investigation

Article Title: Polyamine sequestration of 2 ′ 3 ′ -cGAMP constrains intercellular transmission and STING engagement to subvert antitumor immunity

doi: 10.1172/JCI201460

Figure Lengend Snippet: ( A – C ) IB analysis of lysates from mouse PMs were pretreated with different doses of spermine ( A ), spermidine ( B ), or 10 μM polyamines ( C ) for 1 hour, then incubated with 2′3′-cGAMP-biotin for 4 hours, followed by IP with streptavidin. ( D ) IB analysis of STING dimerization in mouse PMs pretreated with spermine or spermidine, followed by transfection with 2′3′-cGAMP. STING dimerization levels were quantitated by measuring dimer-STING band intensities using ImageJ software; the values were normalized to actin (bar chart). ( E and F ) Molecular docking analysis of spermine ( E ) or spermidine ( F ) interactions with 2′3′-cGAMP, c-di-AMP, c-di-GMP, or 3′3′-cGAMP. ( G – J ) Molecular docking of STING protein with 2′3′-cGAMP ( G ) supplemented with spermine ( H ), spermidine ( I ), and putrescine ( J ). ( K ) Representative confocal images of PLA between spermine or spermidine and STING in PMs pretreated with spermine or spermidine and transfected with 2′3′-cGAMP for 1 hour. The assembly of STING with polyamines was quantitated by the fluorescence intensity using ImageJ software (chart). Scale bars, 10 μm. ( L ) Lysates from mouse PMs pretreated with spermine-, spermidine-, or putrescine-biotin and transfected with 2′3′-cGAMP for 2 hours, followed by IP with streptavidin. IB analysis of STING expression. ( M ) Schematic of spermine and spermidine affecting the binding affinity between 2′3′-cGAMP and STING. Images were created with BioRender. Similar results were obtained from 3 independent experiments. Put, putrescine; Spm, spermine; Spd, spermidine; Trans., transfection; US, unstimulated.

Article Snippet: 2′3′-cGAMP (catalog tlrl-nacga23), 3′3′-cGAMP (catalog tlrl-nacga), c-di-AMP (catalog tlrl-nacda), c-di-GMP (catalog tlrl-nacdg), poly(dG:dC) (catalog tlrl-pgcn), and G3-YSD (catalog tlrl-ydna) were purchased from InvivoGen.

Techniques: Incubation, Transfection, Software, Fluorescence, Expressing, Binding Assay