Journal: Medical mycology

Article Title: A fucose specific lectin from Aspergillus flavus induced interleukin-8 expression is mediated by mitogen activated protein kinase p38.

doi: 10.1093/mmy/myw066

Figure Lengend Snippet: Figure 3. Interleukin-8 expression profile upon FFL treatment: Cells were treated with indicated concentrations of FFL for different time period. Total RNA was isolated and cDNA was synthesized as mentioned, mRNA expression level was determined by Q-PCR method. (A) Cell viability assay: Cells were treated with varying concentrations of FFL for 24 hours. Cell viability was determined as explained in methods. The untreated cells were considered as 100% viable. (B) Cell number assay: Cells were stimulated with varying concentrations of FFL for 24 hours. The cell number in untreated sample was taken as 100%. Trypan blue exclusion assay was used to count the cells. (C) Time dependent expression of IL-8 mRNA in L-132 cells. Cells without treatment were used as control. (D) Dose dependent expression of IL-8 mRNA in L-132 cells. (E) Inhibition of IL-8 mRNA expression by L-fucose: The 40 ng/ml of FFL was preincubated with different concentrations of L-fucose (0.1 mM, 0.2 mM, and 0.3 mM) for 1 hour prior treatment to the cells for indicated time. RNA was isolated and mRNA expression level was determined as explained earlier. (F, G) Expression of IL-8 mRNA in L-132, U-937, and PBMCs: Cells were treated with FFL for 6 and 12 hour. The mRNA expression was determined as explained earlier. (H) Measurements of IL-8 production by sandwich ELISA in L-132, U-937, and PBMCs: Cells were treated with 40 ng/ml of FFL for indicated time. After each time course supernatant was collected and IL-8 concentration was determined as mentioned in kit. The cells treated with 100 nM TPA for 4 hours was used as positive control and untreated cells considered as negative control. The data are presented as mean ± SD of three individual experiments that gave similar results. ∗∗P < .01, ∗∗∗P < .001 versus control. #P < .05, ##P < .01, ###P < .001 versus FFL (40 ng/ml).

Article Snippet: Enzyme-linked immunosorbent assay (ELISA) Interleukin-8 concentration in cell culture supernatant was determined by sandwich ELISA using human IL-8 ELISA kit from Elabscience, Beijing (E-EL-H0048), following manufacturers protocol.

Techniques: Expressing, Isolation, Synthesized, Viability Assay, Trypan Blue Exclusion Assay, Control, Inhibition, Sandwich ELISA, Concentration Assay, Positive Control, Negative Control

Journal: Frontiers in Microbiology

Article Title: Probing Clostridium difficile Infection in Complex Human Gut Cellular Models

doi: 10.3389/fmicb.2019.00879

Figure Lengend Snippet: Production of C. difficile spores, toxins, and host responses to infection. (A) Colony counts of spores recovered after heat treatment, and total cells in the cell-associated C. difficile fraction (infected cell lysates) in the 2D epithelial model. Data shown are the mean of three independent experiments and error bars indicate SD, * p < 0.05 as determined by two-way ANOVA. (B) ELISA for C. difficile toxins A and B shows increased toxin production after extended infection. Toxins were measured from medium obtained from the apical compartment containing uninfected cell layers incubated for 24 or 48 h (Control) or cells infected with C. difficile for 3, 6, 24, or 48 h. Data shown are the mean of three independent experiments and error bars indicate SD, * p < 0.05 as determined by two-way ANOVA. Gray line represents the sensitivity of the test at 0.5 ng/ml. (C) ELISA for human IL-8 indicates increased IL-8 production at 24 and 48 h p.i. IL-8 was measured in medium obtained from the basolateral compartment containing uninfected cell layers incubated for 3, 6, 24, or 48 h (Control) or cells infected with C. difficile for 3–48 h. Gray line represents the limit of detection at 32 pg/ml. Data shown are the mean of three independent experiments and error bars indicate SD, * p < 0.05, ** p < 0.01, and *** p < 0.001, **** p < 0.0001 as determined by one-way ANOVA with Tukey’s test for multiple comparison.

Article Snippet: IL-8 production was also determined by analysis of basolateral supernatants from the VDC using a human IL-8 ELISA kit (R&D systems, Minneapolis, MN, United States) following the manufacturer’s instruction.

Techniques: Infection, Enzyme-linked Immunosorbent Assay, Incubation, Control, Comparison

Journal: Frontiers in Microbiology

Article Title: Probing Clostridium difficile Infection in Complex Human Gut Cellular Models

doi: 10.3389/fmicb.2019.00879

Figure Lengend Snippet: C. difficile spores and toxin production, and host response to infection in the 3D-VDC model. (A) Colony counts of spores and total cells in the host cell-associated C. difficile fraction (infected cell lysates). Data shown are the mean of three independent experiments and error bars indicate SD, ns, not significant as determined by two-way ANOVA. (B) Toxin A and B levels from apical compartment supernatants as determined by ELISA in the 3D model. Data shown are the mean of three independent experiments and error bars indicate SD, ns, not significant as determined by two-way ANOVA. Gray line represents the sensitivity of the test at 0.5 ng/ml. (C) Human IL-8 levels in supernatants from the basolateral compartments with uninfected cells incubated for 24 h or with cells infected with C. difficile for 3 and 24 h, as determined by ELISA. Gray line represents the limit of detection at 32 pg/ml. Data shown are the mean of three independent experiments and error bars indicate SD, ** p < 0.01, as determined by the one-way ANOVA with Tukey’s test for multiple comparison.

Article Snippet: IL-8 production was also determined by analysis of basolateral supernatants from the VDC using a human IL-8 ELISA kit (R&D systems, Minneapolis, MN, United States) following the manufacturer’s instruction.

Techniques: Infection, Enzyme-linked Immunosorbent Assay, Incubation, Comparison

Journal: Cancer Immunology, Immunotherapy

Article Title: Micro-environmental cross-talk in an organotypic human melanoma-in-skin model directs M2-like monocyte differentiation via IL-10

doi: 10.1007/s00262-020-02626-4

Figure Lengend Snippet: Cytokine release by the melanoma reconstructed human skin (Mel-RhS) compared to its control (RhS). After 4 weeks culture at the air–liquid interface, medium was refreshed and culture supernatant was collected over a period of 24 h. Cytokines were detected by means of ELISA. Results are shown as mean ± SEM (* p < 0.05, ** p < 0.01, and *** p < 0.001; paired t test; N = 7 independent experiments performed in duplicate for CCL2, CCL22, IL-8, and GM-CSF; N = 9 independent experiments performed in duplicate for CXCL10, IL-10, and VEGF; N = 10 independent experiments performed in duplicate for M-CSF and TGF-β1)

Article Snippet: Interleukin-8 (IL-8) and IL-10 were measured using human IL-8 ELISA kit (Sanquin, Amsterdam, The Netherlands) and human IL-10 ELISA kit (Diaclone SAS, Besançon, France), respectively.

Techniques: Control, Enzyme-linked Immunosorbent Assay

Journal: Frontiers in Cellular and Infection Microbiology

Article Title: Interkingdom signaling elicited by bacterial extracellular vesicles in human cystic fibrosis airway epithelium and neutrophils

doi: 10.3389/fcimb.2026.1695102

Figure Lengend Snippet: P. aeruginosa EVs are enriched in metabolic pathways and secretory, which can be modified to alter epithelial airway secretion in CF. (A) EVs from bacterial clinical strains were isolated and subjected to MS-based proteomics. The spectra were searched using the MaxQuant algorithm against a reviewed “ P. aeruginosa ” protein sequence database (UniProt). (A) The identified protein was analyzed using GO enrichment analysis performed on the GO home page ( https://geneontology.org/docs/go-enrichment-analysis ) connected to the Panther classification system. The top 10 enriched biological processes from P. aeruginosa are listed in a pie chart. (B) A table of 15 P. aeruginosa EV proteins. (C) EVs were isolated from WT (PAO1) and mutated strains (−OprF, −ADC, −ADI) of P. aeruginosa . The resulting secretion from apical EV and basal cultures was measured using NTA. The number of particles per mL of EVs in airway cultures incubated with WT P. aeruginosa EVs is displayed in [ (C) (i)]. The number of particles per mL of EVs in airway cultures incubated with P. aeruginosa EVs derived from mutated strains with deletions in the following genes is displayed by bar chart ( n = 3) with OprF displayed in [ (C) (ii)], arginine deaminase (ADI−) in [ (C) (iii)], and arginine decarboxylase (ADC−) in [ (C) (iv)]. (D) The levels of IL-8 secretion under the above conditions were analyzed by ELISA, displayed as bar charts. Non-parametric Kruskal–Wallis statistical tests were used for all the above analyses featuring three or more groups.

Article Snippet: Clear supernatants were diluted 1:10, and IL-8 was quantified using the AuthentiKineTM human IL-8 ELISA kit (Proteintech, Chicago, IL, United States) following the manufacturer’s protocol.

Techniques: Modification, Isolation, Sequencing, Incubation, Derivative Assay, Enzyme-linked Immunosorbent Assay

Journal: Scientific Reports

Article Title: Metapanax delavayi extract as a neurocutaneous modulator via CRHR1/POMC/MC1R signaling

doi: 10.1038/s41598-026-39343-4

Figure Lengend Snippet: MDE suppressed substance P-induced release of inflammatory factors in HaCaT cells. ( A ) CCK-8 assay, HaCaT cells were supplemented with 15, 30, 60, 120, 240, 480 μg/mL MDE for 24 h ( N = 3). ( B - D ) RT-qPCR was used to determine the mRNA expression levels of proinflammatory cytokines ( IL-6 , IL-8 , and IL-1α ) ( N = 4). ( E - G ) The content of IL-6, IL-8 and IL-1α in different groups ( N = 5). Data are presented as mean ± SEM. p value using the one-way ANOVA. * p ≤ 0.05, ** p ≤ 0.01 versus control; # p ≤ 0.05, ## p ≤ 0.01 versus substance P-induced.

Article Snippet: The expression of cortisol (Catalogue Numbers: CEA806Mu) (Cloud-Clone Corp, China), β-endorphin (Catalogue Numbers: CSB-E06821h), IL-6 (Catalogue Numbers: CSB-E04638h), IL-8 (Catalogue Numbers: CSB-E04641h), and IL-1α (Catalogue Numbers: CSB-E04620h) (all from Cusabio, China) were also quantified using ELISA kits according to the manufacturer’s instructions.

Techniques: CCK-8 Assay, Quantitative RT-PCR, Expressing, Control