human pro collagen i alpha 1 col1a1 duoset elisa  (R&D Systems)

Journal: npj Biomedical Innovations

Article Title: Developing a protocol to counteract spontaneous in vitro activation of intestinal fibroblasts using design of experiments

doi: 10.1038/s44385-026-00079-5

Figure Lengend Snippet: A Immunofluorescence staining of the cell cytoplasm using Cell Mask (green) and Phalloidin (red). B , C Quantification of cell area and aspect ratio. Gene expression analysis of D ACTA2 , E MYLK , and F MYH11 via qRT-PCR. G Immunofluorescence staining of α-SMA (red) and FAP (green), with cell nuclei counterstained with DAPI (blue). Quantification of the staining intensity of the myofibroblastic protein markers H α-SMA, I FAP. Gene expression analysis of J COL3A1 and K COL1A1 via qRT-PCR. L Quantification of secreted collagen I using ELISA. Data are presented as mean ± SD ( n = 3–5 replicates). Scale bar indicates 100 μm. One-way ANOVA followed by Tukey’s multiple comparisons test. * p < 0.05, ** p < 0.01,*** p < 0.001 and **** p < 0.0001 compared to the control and ## p < 0.01, ### p < 0.001, and #### p < 0.0001 compared to TGF-β 1 .

Article Snippet: Fibronectin and collagen secretion were quantified using commercially available Human Fibronectin (FN) ELISA Kit (RayBiotech, product # ELH-FN1-1) and Human Pro-Collagen I alpha 1 (COL1A1) DuoSet ELISA (R&D Systems, product # DY6220-05) following the manufacturer’s instructions.

Techniques: Immunofluorescence, Staining, Gene Expression, Quantitative RT-PCR, Enzyme-linked Immunosorbent Assay, Control

Journal: npj Biomedical Innovations

Article Title: Developing a protocol to counteract spontaneous in vitro activation of intestinal fibroblasts using design of experiments

doi: 10.1038/s44385-026-00079-5

Figure Lengend Snippet: A Quantification of cell viability. Gene expression analysis of B ACTA2 , C MYLK , and D MYH11 via qRT-PCR. E Immunofluorescence staining of α-SMA (red) and FAP (green), with cell nuclei counterstained with DAPI (blue). Quantification of the staining intensity of the myofibroblastic protein markers F α-SMA and G FAP. Gene expression analysis of H COL3A1 and I COL1A1 via qRT-PCR. Quantification of ECM protein secretion for J collagen I and K fibronectin using ELISA. Data are presented as mean ± SD ( n = 3–5 replicates). Scale bar indicates 100 μm. One-way ANOVA followed by Tukey’s multiple comparisons test. * p < 0.05, ** p < 0.01, *** p < 0.001 and **** p < 0.0001 compared to the control and # p < 0.05, ## p < 0.01, #### p < 0.0001 compared to TGF β 1 -treated group, for comparisons shown.

Article Snippet: Fibronectin and collagen secretion were quantified using commercially available Human Fibronectin (FN) ELISA Kit (RayBiotech, product # ELH-FN1-1) and Human Pro-Collagen I alpha 1 (COL1A1) DuoSet ELISA (R&D Systems, product # DY6220-05) following the manufacturer’s instructions.

Techniques: Gene Expression, Quantitative RT-PCR, Immunofluorescence, Staining, Enzyme-linked Immunosorbent Assay, Control

Journal: ACR Open Rheumatology

Article Title: Lymphocyte Activation Gene 3 Regulation of Profibrotic Cytokines and Type I Collagen Production in Patients With Systemic Sclerosis

doi: 10.1002/acr2.70120

Figure Lengend Snippet: Impact of agonistic LAG‐3 antibody on proinflammatory cytokine secretion by PBMCs. (A) dcSSc (n = 8) PBMCs were prestimulated with CD3/CD28 for 24 hours in monocultures or (B) cocultured with allogeneic dcSSc fibroblasts. The cultures were treated with either an LAG‐3 Q22 agonistic antibody (LAG‐3 Ag) or an isotype control (LAG‐3 Isotype) for 48 hours. In monocultures, the agonistic LAG‐3 Ag significantly reduced the levels of IFNγ, IL‐1β, IL‐4, and TNFα. In cocultures, IFNγ, IL‐12p70, IL‐13, IL‐2, IL‐4, and TNFα were decreased significantly. In both setups, IL‐10 levels were significantly increased by the addition of agonistic LAG‐3 isotype–treated samples. Data are presented as mean with SD Q23 (* P < 0.05, ** P < 0.01). Ag, agonistic antibody; dcSSc, diffuse cutaneous systemic sclerosis; IFNγ, interferon γ; IL, interleukin; LAG‐3, lymphocyte Q24 activation gene 3; ns, not significant; NT, no treatment; PBMC, peripheral blood mononuclear cell; TNFα, tumor necrosis factor α (1425 pg/mL vs 1847 pg/mL; P = 0.001); compared to the control IgG, there was still a notable increase in IL‐10 production (8.33 pg/mL vs 6.10 pg/mL; P = 0.01). No significant changes were observed in the production of IL‐1β or IL‐6 (Figure ). Agonistic LAG‐3 antibodies suppress type I IFN production and reduce fibroblast matrix production. PBMCs from patients with dcSSc (n = 8) were stimulated with CD3/CD28 for 24 hours and cultured with or without allogeneic dcSSc fibroblasts for 48 hours. The cultures were then treated with either the LAG‐3 Ag or the isotype control (LAG‐3 Isotype). (C, D) The LAG‐3 Ag significantly reduced the production of bioactive IFNα/β in monocultures and cocultures, as measured in HEK‐Blue cells, compared to the LAG‐3 isotype control. (E) In cocultures, levels of type 1 procollagen and (F) fibronectin were significantly reduced by LAG‐3 Ag. Data are presented as mean with SD (* P < 0.05, ** P < 0.01). Ag, agonistic antibody; dcSSc, diffuse cutaneous systemic sclerosis; IFN, interferon; LAG‐3, lymphocyte activation gene 3; ns, not significant; NT, no treatment; PBMC, peripheral blood mononuclear cell.

Article Snippet: Quantification of sLAG‐3 (LAG‐3 Human Enzyme‐Linked Immunosorbent Assay [ELISA] kit # BMS2211; Invitrogen), type 1 procollagenα (Human Pro‐Collagen I alpha 1 DuoSet ELISA Cat.DY6220‐05; R&D Systems), and fibronectin (Human Fibronectin DuoSet ELISA Cat. DY1918‐05; R&D Systems) in the plasma and supernatants was performed according to the manufacturer's protocol.

Techniques: Control, Activation Assay, Cell Culture