Journal: Cancers

Article Title: Development of Ex Vivo Analysis for Examining Cell Composition, Immunological Landscape, Tumor and Immune Related Markers in Non-Small-Cell Lung Cancer

doi: 10.3390/cancers16162886

Figure Lengend Snippet: Expression of lung cancer markers and immune cell composition in surgically resected NSCLC and lung tissue samples analyzed on the ex vivo cell preparations.

Article Snippet: After washing with PBS, the other ex vivo cell preparations were permeabilized with 0.3% Triton X-100 solution in PBS for 2 min, blocked in PBS solution containing 2% BSA, and incubated with the appropriate rabbit or mouse primary antibodies to human cancer cell markers: TTF1 (clone SP141, Abcam, Cambridge, UK, ab227652), p40 (clone EPR17863-47, Abcam, Cambridge, UK, ab203826), CK7 (clone PBM-12F1, PrimeBioMed, Moscow, Russia), PanCK, including the cytokeratin 5/6 (clone C11, PrimeBioMed, Moscow, Russia), Ki-67 (clone SP6, Thermo Fisher Scientific, Waltham, MA, USA, MA5-14520), AhR (clone RPT9, Abcam, Cambridge, UK, ab2769), AhRR (Abcam, Cambridge, UK, ab234817), CYP1A1 (MyBioSource, San Diego, CA, USA, MBS178240); to immune-related markers: PD-L1 (clone ABM4E54, Abcam, Cambridge, UK, ab210931), PD-1 (clone CAL20, Abcam, Cambridge, UK, ab237728), CD14 (clone SP192, Spring Bioscience, Pleasanton, CA, USA, M492), IFNγ (clone 25718, Thermo Fisher Scientific, Waltham, MA, USA, MA5-23718), TNFα (Thermo Fisher Scientific, Waltham, MA, USA, P300A), FGFb (clone 6/basic FGF, BD Biosciences, Franklin Lakes, NJ, USA, 610072), IL-1β (clone 2805, Thermo Fisher Scientific, Waltham, MA, USA, MA5-23691), IL-4 (Thermo Fisher Scientific, Waltham, MA, USA, PA5-25165), IL-10 (clone 945A2A5, Invitrogen, Waltham, MA, USA, AHC9102), IL-12 (clone 24945, Thermo Fisher Scientific, Waltham, MA, USA, MA5-23715), and to fibroblast markers: CD10 (clone GM-003, PrimeBioMed, Moscow, Russia), a membrane metalloendopeptidase [ ], and TRITC-labeled phalloidin dye (Sigma-Aldrich, Burlington, MA, USA, P1951) to stain filamentous actin (F-actin), diluted 1:100 each.

Techniques: Expressing, Ex Vivo, Marker