pathscan human phospho slp 76 ser376 sandwich 384 well elisa kits  (Cell Signaling Technology Inc)

Journal: iScience

Article Title: Staphylococcus aureus induced trained immunity in macrophages confers heterologous protection against gram-negative bacterial infection

doi: 10.1016/j.isci.2024.111284

Figure Lengend Snippet: S. aureus exposure enhances human macrophage responses to subsequent LPS or E. coli challenge (A and B) Human peripheral blood monocytes were isolated by CD14 + selection and exposed to heat-killed S. aureus strain Newman (2 μg/ml) or PBS. Media was refreshed after 24 h and cells were maintained in RPMI supplemented with 10% pooled human serum. On day 7 macrophages were subsequently exposed to LPS and cytokine responses measured at the mRNA level at 3 h by RT-PCR (A) and protein level at 24 h by ELISA (B). (C) PBS and S. aureus exposed macrophages were also rechallenged with live E. coli (MOI100) and cytokine responses measured by ELISA at 24 h post challenge. Results expressed pg/ml or ΔΔct +/−SEM for n = 6–8 individual donors. Statistical significance measured by paired t test, p < 0.05 ∗, p < 0.01 ∗∗, p < 0.0001 ∗∗∗.

Article Snippet: Human H3 acetylation ELISA , Cell Signaling Technology , 7232.

Techniques: Isolation, Selection, Reverse Transcription Polymerase Chain Reaction, Enzyme-linked Immunosorbent Assay

Journal: iScience

Article Title: Staphylococcus aureus induced trained immunity in macrophages confers heterologous protection against gram-negative bacterial infection

doi: 10.1016/j.isci.2024.111284

Figure Lengend Snippet: S. aureus exposure leads to histone modifications in macrophages that are required to support enhanced responses to subsequent LPS challenge (A and B) Human peripheral blood monocytes were isolated by CD14 + selection and exposed to heat-killed S. aureus strain Newman (2 μg/ml) or PBS. Macrophage lysates were collected on day 3 post training and sonicated to shear DNA. To assess histone 3 acetylation a pathscan ELISA on lysates was carried out (A). Monocytes were exposed to heat-killed S. aureus strain Newman (2 μg/ml) or PBS in the presence or absence of the methyltransferase inhibitor MTA (1 mM). Media containing MTA and the training stimulus was removed after 24 h and replaced with RPMI supplemented with 10% pooled human serum and stimulated with LPS (10 ng/ml) on day 7. IL-6, TNF, and IL-10 production was measured by ELISA (B). Results expressed as pg/ml or absorbance at 450 ± SEM for n = 5–9 individual donors. Statistical significance measured by paired t test, p < 0.05 ∗, p < 0.01 ∗∗.

Article Snippet: Human H3 acetylation ELISA , Cell Signaling Technology , 7232.

Techniques: Isolation, Selection, Sonication, Shear, Enzyme-linked Immunosorbent Assay

Journal: iScience

Article Title: Staphylococcus aureus induced trained immunity in macrophages confers heterologous protection against gram-negative bacterial infection

doi: 10.1016/j.isci.2024.111284

Figure Lengend Snippet: S. aureus exposure in mice offers protection upon subsequent E. coli challenge (A–D) Groups of C57/Bl6 mice were exposed to S. aureus through i.p. injection (5 × 10 8 cfu/ml) on days 0, 7, and 14. Mice were allowed to rest for 21 days upon which mice are rechallenged i.p. with E. coli (1.5 × 10 7 cfu/ml). Proportions of Ly6G + CD11b+ neutrophils and Ly6G-/CD11b+/Ly6C+ monocytes were analyzed by flow cytometry at 3 h post E. coli challenge (A) and expression of MPO and iNOS (MFI) on monocytes assessed by flow cytometry at 3 h post E. coli challenge (B). IL-6, TNF, and IL-1β levels in the peritoneal lavage fluid were quantified by ELISA at 3h post E. coli challenge (C). At 3 h and 24 h post E. coli challenge peritoneal cavity was lavaged and kidneys, lungs, livers were excised and homogenized. Lavage and homogenates were plated on TSA and allowed to grow overnight and total bacterial burden assessed (D). Results expressed as % of cells, mean fluorescence Intensity, pg/ml or LOG10CFU per mL +/−SEM for n = 4–10 individual mice. Statistical significance measured by unpaired t test (A–C) or two-way ANOVA with Sidak’s test for multiple comparisons (D), p < 0.05 ∗, p < 0.01 ∗∗, p < 0.0001 ∗∗∗.

Article Snippet: Human H3 acetylation ELISA , Cell Signaling Technology , 7232.

Techniques: Injection, Flow Cytometry, Expressing, Enzyme-linked Immunosorbent Assay, Fluorescence

Journal: iScience

Article Title: Staphylococcus aureus induced trained immunity in macrophages confers heterologous protection against gram-negative bacterial infection

doi: 10.1016/j.isci.2024.111284

Figure Lengend Snippet: S. aureus exposure enhances myelopoiesis (A) Groups of C57Bl6 mice received a single i.p. challenge of S. aureus (5 × 10 8 cfu). 3 h post challenge blood was collected and the left femur was excised. Femurs were flushed with 500 μl sterile PBS, cells were removed by centrifugation and 100 μl of the extracellular fluid and blood was plated on TSA and allowed to grow overnight and total bacterial burden assessed. (B) IL-1 β levels in bone marrow extra-cellular fluid was assessed by ELISA at 3 h post S. aureus challenge. (C–F) Groups of C57/Bl6 mice were exposed to S. aureus through i.p. injection (5 × 10 8 cfu/ml) on days 0, 7, and 14. Mice were allowed to rest for 21 days upon which, bone marrow cells were isolated for phenotypic analysis by flow cytometry. Proportion of LKS cells (Lineage - cKit + Sca1 + ) (C), LT-HSCs (LKS + CD48 − CD150 + ) (D), ST-HSCs (LKS + CD48 + CD150 + ) (E), and MPP cells (LKS+CD48 + CD150-) (F) in bone marrow were assessed. (G) Proportion of MPP3 (LKS+CD48 + CD150-Flt3+) and MPP4 (LKS+CD48 + CD150-Flt3-) cells were also assessed. (H and I) Neutrophil (H) and monocyte (I) numbers were assessed in bone marrow and in the blood on day 21 post final S. aureus or PBS exposure ( n = 5–6). Results expressed as % of cells, absolute cell number, pg/ml or CFU per mL +/−SEM. Statistical significance measured by unpaired t test, p < 0.05 ∗, p < 0.01 ∗∗, p < 0.0001 ∗∗∗.

Article Snippet: Human H3 acetylation ELISA , Cell Signaling Technology , 7232.

Techniques: Sterility, Centrifugation, Enzyme-linked Immunosorbent Assay, Injection, Isolation, Flow Cytometry

Journal: iScience

Article Title: Staphylococcus aureus induced trained immunity in macrophages confers heterologous protection against gram-negative bacterial infection

doi: 10.1016/j.isci.2024.111284

Figure Lengend Snippet: S. aureus exposure enhances macrophage effector functions Groups of C57/Bl6 mice were exposed to S. aureus or PBS through i.p. injection (5 × 10 8 cfu/ml) on days 0, 7, and 14. Mice were allowed to rest for 21 days upon which BMDMs were generated and then challenged in vitro with LPS (10 ng/ml) or E. coli (MOI 100). (A–D) IL-6 and TNF production was measured by ELISA at 24 h (A). H3K27ac was assessed in BMDMs lysates from PBS and S. aureus exposed mice by ELISA (B). BMDMs from S. aureus exposed or PBS exposed mice were adoptively transferred into naive recipient mice 4 h prior to E. coli (1.5 × 10 7 cfu) challenge. At 3 h post E. coli challenge, the peritoneal cavity was lavaged and the proportions of CD11b+/Ly6G + neutrophils analyzed by flow cytometry (C). At 3 h and 24 h post E. coli challenge, peritoneal lavage was collected and the lung, liver, and kidney were excised. Organs were homogenized and plated along with peritoneal lavage on TSA and allowed to grow overnight and total bacterial burden assessed (D). Results expressed as % of cells, pg/ml, absorbance at 450, LOG10CFU per mL for n = 6–10 individual mice, +/−SEM. Statistical significance measured by unpaired t test (A–C) or two-way ANOVA with Sidak’s test for multiple comparisons (D), p < 0.05 ∗, p < 0.01 ∗∗, p < 0.0001 ∗∗∗.

Article Snippet: Human H3 acetylation ELISA , Cell Signaling Technology , 7232.

Techniques: Injection, Generated, In Vitro, Enzyme-linked Immunosorbent Assay, Flow Cytometry

Journal: iScience

Article Title: Staphylococcus aureus induced trained immunity in macrophages confers heterologous protection against gram-negative bacterial infection

doi: 10.1016/j.isci.2024.111284

Figure Lengend Snippet:

Article Snippet: Human H3 acetylation ELISA , Cell Signaling Technology , 7232.

Techniques: Control, Virus, Enzyme-linked Immunosorbent Assay, Software, Flow Cytometry

Journal: Integrative biology : quantitative biosciences from nano to macro

Article Title: Modeling Shiga toxin-induced human renal-specific microvascular injury

doi: 10.1093/intbio/zyae001

Figure Lengend Snippet: Antibodies used.

Article Snippet: 15 μ g of protein per lane were electrophoresed on NuPAGE 4–12% Bis-Tris Acrylamide gels (Invitrogen), transferred to nitro-cellulose membrane, blocked with 5% milk in TBS and probed overnight at 4°C with 1:1000 rabbit-anti-human cleaved caspse-3 (Cell Signaling) and 1:1200 mouse-anti-human β -tubulin (Cell Signaling) primary antibodies in 5% milk in TBS-0.1% Tween.

Techniques: Control

Journal: MedComm

Article Title: Co‐targeting FAK and Gli1 inhibits the tumor‐associated macrophages‐released CCL22‐mediated esophageal squamous cell carcinoma malignancy

doi: 10.1002/mco2.381

Figure Lengend Snippet: C‐C motif chemokine 22 (CCL22) promotes esophageal squamous cell carcinoma (ESCC) progression . (A) The secretion of CCL22 from primary tumor‐associated macrophages (PriTAMs) was evaluated using enzyme‐linked immunosorbent assay (ELISA). (B) Protocols of invasion assay. (C) Invasive ability of KYSE410 (upper panel) or KYSE510 (lower panel) cells in the presence of CCL22‐positive PriTAMs with/without CCL22 Ab, or CCL22‐negative PriTAMs or rhCCL22 (50 ng/mL). (D) Soft agar‐based colony formation of KYSE410 (upper panel) or KYSE510 (lower panel) cells treated with conditioned media (CM) from CCL22‐positive PriTAMs with/without CCL22 Ab, or CCL22‐negative PriTAMs, or with rhCCL22 (50 ng/mL). (E) KYSE410 and KYSE510 cells were transfected with CCR4 siRNAs, and the transfection efficiency was evaluated using immunoblotting. Glyceraldehyde‐3‐phosphate dehydrogenase (GAPDH) served as internal control. (F–G) The invasive or anchorage‐independent growth ability of indicated ESCC cells in the presence of rhCCL22 (50 ng/mL) was evaluated using transwell invasion assay (F) or soft agar‐based colony formation assay (G). *** p < 0.001 as compared with the control cells. $$$ p < 0.001 as compared with PriTAMs #1, #2, or #3. Two‐tailed unpaired Student's t ‐test. Error bars, mean ± SD of five independent experiments.

Article Snippet: Human Sox2 ELISA kit (CST; Cat# 7277), human Nanog ELISA kit (Abcam; Cat# ab236720), and human OCT‐4 ELISA kit (Raybiotech; Cat# ELH‐OCT4‐1) were applied to measure expression of stemness‐related biomarkers, including SOX2, Nanog, and OCT4 in indicated ESCC cells.

Techniques: Enzyme-linked Immunosorbent Assay, Invasion Assay, Transfection, Western Blot, Control, Transwell Invasion Assay, Soft Agar Assay, Two Tailed Test

Journal: MedComm

Article Title: Co‐targeting FAK and Gli1 inhibits the tumor‐associated macrophages‐released CCL22‐mediated esophageal squamous cell carcinoma malignancy

doi: 10.1002/mco2.381

Figure Lengend Snippet: Combination of VS‐4718 and GDC‐0449 in esophageal squamous cell carcinoma (ESCC) treatment in vivo . (A) KYSE510 (upper panel) or KYSE410 (lower panel) tumor‐beared mice with (right panel)/without (left panel) rhCCL22 were treated with VS‐4718 or GDC‐0449 and their combination. Curves of tumor volumes were shown. (B–D) The expression of Ki‐67 (B), CD31 (C), or LYVE1 (D) in indicated KYSE410 and KYSE510 tumors was evaluated using quantitative enzyme‐linked immunosorbent assay (ELISA) assays. Black bar: KYSE410 tumor, left panel of KYSE410 group: cont (without C‐C motif chemokine 22 [CCL22]), right panel of KYSE410 group: with CCL22; gray bar: KYSE510 tumor, left panel of KYSE510 group: cont (without CCL22), right panel of KYSE510 group: with CCL22; * p < 0.05; *** p < 0.001 as compared with the control cells. Error bars, mean ± SD of five independent experiments.

Article Snippet: Human Sox2 ELISA kit (CST; Cat# 7277), human Nanog ELISA kit (Abcam; Cat# ab236720), and human OCT‐4 ELISA kit (Raybiotech; Cat# ELH‐OCT4‐1) were applied to measure expression of stemness‐related biomarkers, including SOX2, Nanog, and OCT4 in indicated ESCC cells.

Techniques: In Vivo, Expressing, Enzyme-linked Immunosorbent Assay, Control