anti-mouse integrin α5β1 monoclonal antibody  (Merck & Co)

Journal: Frontiers in Immunology

Article Title: Dynamic roles of neutrophil extracellular traps in cancer cell adhesion and activation of Notch 1-mediated epithelial-to-mesenchymal transition in EGFR-driven lung cancer cells

doi: 10.3389/fimmu.2024.1470620

Figure Lengend Snippet: Cell adhesion of different cancer cell lines in the presence or absence of NETs. Cells were seeded and allowed to adhere to NET-coated multi-well plates for 1, 2 or 4 h depending on the cell line (A–D) . Negative controls were plates coated with PBS or conditioned medium of dHL-60 and NET-coated plates pre-incubated with DNase I. Results are expressed as percentage of total cell number in each well. In parallel experiments, adherent cells were exposed to 0.5 µg/mL NETs for 48 h followed by counting of adherent and detached viable cells (E) . (A) Cell adhesion (mean ± SE) obtained in HCC827 (2 h), H1975 (2 h), H1993 (2 h) and A549 (1 h) lung cancer cell lines. (B) Cell adhesion (mean ± SE) obtained in MCF7 (2 h) and MDA MB231 (2 h) breast cancer cells, HT1080 fibrosarcoma (1 h) and U87-MG glioblastoma (4 h) cell lines. (C) Cell adhesion (mean ± SE) to NET-coated plates obtained in HCC827 cells pre-incubated with blocking antibodies against α5β1 integrin and CCDC25 receptor. (D) Cell adhesion (mean ± SE) to NET-coated plates obtained in H1975 cells pre-incubated with blocking antibodies against α5β1 integrin and CCDC25 receptor. (E) Percentage of adherent and detached viable cells exposed or not to NETs in the medium for 48 h. Statistical significance: *p<0.05; **p< 0.01; ***p< 0.001; ns, not significant.

Article Snippet: To test the role of α5β1 integrin and CCDC25 receptor in promoting cell adhesion to NETs, 3×10 5 cells were pre-incubated with 5μg/ml of blocking antibody recognizing α5β1 integrin (Millipore Cat# MAB2514, RRID: AB_94626) or CCDC25 (Thermo Fisher Scientific Cat# PA5-54735, RRID: AB_2639443) in 300μl of serum-free medium supplemented with 1% BSA for 1 h at 37°C and 5% CO 2 .

Techniques: Incubation, Blocking Assay

Journal: Bioengineering & translational medicine

Article Title: 3D bioprinting of an implantable xeno-free vascularized human skin graft.

doi: 10.1002/btm2.10324

Figure Lengend Snippet: FIGURE 5 Phenotyping characterization of human keratinocytes under xeno-free conditions. (a) Live phasecontrast microscopy images of keratinocytes after isolation from the epidermis of donor foreskin and at the confluency state. (b) The cumulative population doublings of keratinocytes cultured under xeno-free conditions is comparable to KGM-Gold medium. (c) Flow cytometry analysis confirmed the expression of integrins α2β1, α5β1, α6, α3, and β4, but not αvβ3. (d) Confocal microscopy exhibiting CK14, CK10, junctional ZO-1, and intracellular occludin staining. Scale bar = 100 μm. Representative of three independent donors

Article Snippet: ECs, FBs, PCs, and KCs cultured under xeno-free conditions were analyzed for surface and intracellular markers expression by flow cytometry and immunofluorescence microscopy using antibodies against: CD31 (WM59; Biolegend), CD45 (2D1; Biolegend), ZO-1 (sc-33725; Santa Cruz), VE-cadherin (sc-6458; Santa Cruz), vWF (ab201336; abcam) claudin-5 (34-1600; Invitrogen), PDGFR-α (16A1; Biolegend), PDGFR-β (18A2; Biolegend), CD90 (5E10; Biolegend), NG2 (9.2.27; Invitrogen), a-SMA (1A4; Invitrogen), FAP (AF3715; Novus Biologics), integrins α2β1 (ab24697; abcam), α5β1 (NBP2-52680; Novus Biologics), αvβ3 (sc-7312; Santa Cruz), α6 (MAB13501; R&D systems), α3 (MAB1345; R&D systems), β4 (NBP1-43369), CK14 (LL002; Novus Biologics), CK10 (EP1607IHCY, abcam), and occludin.

Techniques: Microscopy, Isolation, Cell Culture, Flow Cytometry, Expressing, Confocal Microscopy, Staining