Journal: Bio-protocol

Article Title: Immunofluorescent Staining of Claudin-2 in Cultured Kidney Tubular Cells

doi: 10.21769/BioProtoc.3678

Figure Lengend Snippet: Phase contrast images of live cells grown on cover glasses were obtained using a Nikon eclipse TS100 microscope (10x objective).

Article Snippet: Micro cover glasses, round, No. 1, 18 mm (VWR ® International, catalog number: CA-48380-046), sterilize coverglasses using autoclave Microscope slides (Fisherbrand, Fisher Scientific, catalog number: 12-550-15) Clear tissue culture-treated 12-well microplates, (Corning Costar ® , Millipore-Sigma, catalog number: CLS3513) Parafilm Paper towel Cell line: LLC-PK 1 kidney tubule epithelial cell line (European Collection of Authenticated Cell Cultures (Salisbury, UK), catalog number: 86121112) For cell culture: Dulbecco's Modified Eagle Medium (D-MEM), low glucose, with L-glutamine 110 mg/L sodium pyruvate (Thermo Fisher, Gibco, catalog number: 11885084) 10% Fetal Bovine Serum (FBS) (Thermo Fisher, Gibco, catalog number: 12483-020) 1% Penicillin-Streptomycin (Penicillin-Streptomycin, 100x, sterile-filtered, cell culture tested) (Millipore-Sigma Aldrich, catalog number: P-4333) Trypsin-EDTA (0.05% Trypsin with EDTA 4Na) (Thermo Fisher, Gibco, catalog number: 25300062) Phosphate buffered saline (PBS) (Thermo Fisher, Gibco, catalog number: 10010023) Methanol (BioShop Canada, catalog number: Met302), cool to -20 °C before using 10x PBS Buffer (pH 7.4 Sterile, w/o Ca, Mg) (BioShop Canada, catalog number: PBS415), open in hood to maintain sterility for storage Bovine serum albumin (BSA) (fraction V, heat shock isolation, > 98%) (BioShop Canada, catalog number: ALB001) Polyclonal rabbit anti-Claudin-2 antibody (Thermo Fisher Scientific, Invitrogen, catalog number: 28530), aliquot to avoid repeated thawing, store at -20 °C Fluorophore-labeled secondary antibody: Donkey anti-Rabbit IgG labelled with Alexa Fluor 488 (green) (Thermo Fischer Scientific, catalog number: A-21206) or Donkey anti-Rabbit IgG labelled with Alexa Fluor 555 (red) (Thermo Fischer Scientific, catalog number: A- A31572) 2,4-diamidino-2-phenylindole (DAPI) (Thermo Fischer Scientific, catalog number: D1306) Make a 5 mg/ml DAPI stock solution by dissolving the contents of one vial (10 mg) in 2 ml of deionized water.

Techniques: Microscopy

Journal: Journal of visualized experiments : JoVE

Article Title: A Mouse Model to Investigate the Role of Cancer-associated Fibroblasts in Tumor Growth

doi: 10.3791/61883

Figure Lengend Snippet: 2. Generate cancer cells for injection

Article Snippet: Aliquot resuspended cells onto new tissue culture plates of the appropriate size with DMEM + 10% FBS (10 mL of media is suitable for a 10 cm tissue culture plate). table ft1 table-wrap mode="anchored" t5 Name of Material/Equipment Company Catalog Number Comments/Description Centrifuge for conical tubes capable of reaching 180 × g Fisher Scientific 14-432-22 Sterle 25 ml serological pipet Celltreat 667225B Sterile 10 ml serological pipet Celltreat 667210B Sterile 5 ml serological pipet Celltreat 229005B Sterile 50 ml centrifuge tubes Genesee Scientific 28-108 TC treated 150 × 20 mm dishes Genesee Scientific 25-203 TC treated 100 × 20 mm dishes Genesee Scientific 25-202 TC treated 60 × 15 mm dishes Genesee Scientific 25-260 Cell Culture Multi Flasks Fisher Scientific 14-826-95 Dulbecco’s Modified Eagle Medium Fisher Scientific 11965-118 Sterile tissue-culture grade PBS Fisher Scientific 50-751-7476 Sterile tissue culture-grade Trypsin-EDTA Fisher Scientific 15400054 Fetal bovine serum Fisher Scientific MT35010CV Countess Cell Counting Chamber Fisher Scientific C10228 Cancer cells ATCC ATCC® CRL-11147™ This is the catalog number for a primary human melanoma cell line.

Techniques: Cell Culture, Modification, Cell Counting, Derivative Assay, Injection