CORNING Life Sciences

Transcription

1 CORNING Life Sciences Cell Culture Treatments

2 Corning A brief istory - A brief history - Corning s Brands - Corning s Workflows - Cell Culture Workflow - Cell Culture surfaces - Suspension cells - Non treated - ULA = Ultra Low Attachment - Adherent Cells - TC = standard treated culture - Synthetic non specific - Biological coating - Syntethic defined - Syntehtic specific - Other products with specific cell culture surfaces - Inserts cell culture - Assays microplates 2

3 Corning A brief istory Founded in 1851 : world leader in specialty glass and ceramics. Corning created and make keystone components that enable high-technology systems for consumer electronics, mobile emissions control, telecommunications and life sciences : bubble (first one it Edison ; touchable screens ; eadquarters : Corning, New York Employees: Approximately 25,000 worldwide Corning acquired Costar Corporation to become Corning Costar in Corning acquired Axygen end of 2009 Corning acquired Gosselin end of 2010 Corning acquired Falcon DL end of

4 Corning Life Sciences Different Brands & Market segments Whatever is your application, you can now get a more comprehensive line of lab research tools with Corning s quality. FALCON Cell Culture, Drug Discovery & Liquid andling Molecular Biology Genomics Microbiology cellgro is available in Europe as Mediatech. 4

5 Corning Life Science - Workflows 5

6 Corning Life Science - Workflows Cell Culture surfaces : - Suspension cells - Adherent cells Adherent Cells : - Standard - TC - Synthetic improved (CellBIND ; Primaria) - Biological (ECM Biocoat) - Synthetic defined (Purecoat) - Synthetic specific (OsteoAssays / Synthemax) 6

7 Corning Life Science Cell Culture surfaces Adherent culture Moving Roller bottles Microcarriers in spinner flasks Flasks on rockers Stationary Flasks Microplates Dishes Transwell inserts YPERFlask CellSTACK Suspension culture Moving Spinner flasks Shaker flasks Roller bottles Bags Stationary Flasks Storage bottles Dishes Microplates Transwell inserts Bags 7

8 Corning Life Science Cell Culture surfaces Adherent culture Suspension culture Moving 8

9 Corning Life Science Cell Culture surfaces Suspension Cells Suspension Cell Culture : NT (Not Treated) For standard suspension cell culture ULA (Ultra Low Attachment) For advanced suspension cell applications (stem cells..) 9

10 Corning Life Science Cell Culture surfaces Adherent Cells Adherent Cell Culture : Animal Free Specificity to Cells YES - Standard TC - Synthetic improved (CellBIND ; Primaria) YES - Biological (ECM Biocoat) NO - Synthetic defined (Purecoat) YES - Synthetic specific (OsteoAssays / Synthemax) YES 10

11 Standard Tissue Culture Surfaces Standard Tissue Culture (TC) Treatement : To be used with adherent cells, the Polystyrene needs to be treated to add charges and to get it hydrophylic. Polystyrene is fully hydrophobic and no natural. TC-surface treatment is achieved by (similar proceedure) : By gas plasma : placing polystyrene vessels into a vacuum chamber that receives a mixture of gases with electric power. By corona discharge. The treatment will create a modification of the polystyrene altering it s surface chemistry Negative charges Feature No specific 11

12 Standard Tissue Culture Surfaces Culture Vessels - uncharged - hydrophobic - No cell attachment Corona Discharge Plate igh energy oxygen ions Conveyor belt e.g. "non-treated" rounder 40 ml droplet C C C C C C C C C Surface treatment C C C C C C O C O C O O ydrophilic O ydrophobic - charged groups - hydrophilic - cell attachment O C O flatter e.g. "TC" 40 ml droplet 12

13 Synthetic Non Specific CellBIND & Primaria Syntethic Non specific (Improved attachment) CellBIND or Primaria : To be used with adherent cells, the Polystyrene needs to be treated to add charges and to get it hydrophylic. Polystyrene is fully hydrophobic and no natural. TC-surface treatment is achieved by : a specific gaz mixture. A difference source of energy (microwaves). The treatment will create a modification of the polystyrene altering it s surface chemistry more balanced vs + positives Feature No specific ; more dedicated to difficult cells 13

14 Synthetic Non Specific CellBIND & Primaria CellBIND TCT BD Primaria Treatment N2O O2 N4 /proprietary % Oxygen/ Nitrogen 35% Oxygen 22% Oxygen 14.1% Oxygen 11.1% Nitrogen 1.8% Oxygen Contact angle Charge Negative Negative Positive Not Treated TCT CellBIND 14

15 ECM Biological Coating Biological coating with ECM molecules : Cells are not living in 2D into a plastic box! They are normally living in the context of other cells, surrounded and in contact with an extracellular matrix (ECM) The TC-treated surfaces can be coated with ECM components : Biological molecules from the ECM : Can be bought pre-coated ready-to-use products. Can be bought as a chemical to plate into the lab. Reconstituted ECM = Corning Matrigel Matrix : Can be bought pre-coated ready-to-use products. Can be bought as a chemical to plate into the lab. 15

16 ECM Biological Coating READY-TO-USE Biological molecules from the ECM : Collagen I Collagen IV Fibronectin Gelatin Laminin Poly-Lysine Poly-D-Lysine/Laminin Poly-L-Ornithine/Laminin Laminin/Fibronectin Reconstituted ECM = Corning Matrigel Matrix : Matrigel Matrigel Growth Factors Reduced (GFR) 16

17 ECM Biological Coating TO COAT INTO TE LAB : Biological molecules from the ECM : Collagen I, II, III, IV, V, VI Fibronectin Laminin Osteopontin Poly-Lysine Vitronectin Reconstituted ECM = Corning Matrigel Matrix : Matrigel Matrigel hesc (human Embryonic Stem Cells) qualified Matrigel C to do tumor injection Matrigel Phenol Red free Matrigel GFR (Growth Factor Reduced) Dispase & Cell Recovery Solution to recover cells from Matrigel 17

18 ECM Biological Coating APPLICATIONS : Endothelial cell migration, invasion Angiogenesis tube formation Tumor invasion 3-D-cell culture Maintenance of human embryonic stem cells Neurite outgrowth epatocyte culture SEM s of hepatocytes cultured for two days on collagen I (1), collagen I gels (2), or BD Matrigel Matrix (3). Note the cluster of spherical cells for hepatocytes cultured on Corning Matrigel Matrix, typical of differentiated cells. 18

19 ECM Biological Coating APPLICATIONS : EK293 Before Wash After Wash After Wash, Calcein AM Staining After Wash After Wash, Calcein AM Staining TC Plates Before Wash BD BioCoat PDL Plates EK-293 cells were cultured under serum-free conditions on TC-treated or BD BioCoat PDL 384-well black/clear plates. Samples were washed using a Skatron Washer (Molecular Devices). 19

20 ECM Biological Coating ADVANTAGES : More in vivo like properties Better way of life : attachment, proliferation, growth including difficult cells as : primary cells, neural cells, hepatocytes, endothelial cells, Differentiation and functional studies possible 3-dimensional culture of cells possible, when cells are embedded in a thick gel, and not only applied to a thin coating Testing packs to try different molecules (mix of different surfaces) LIMITATION : Not Animal components free Limit shelf life 20

21 Synthetic defined - Purecoat An advanced cell culture surfaces, that uses a novel process to coat the culture vessel in a very defined and reproducible way 2 options : amine (+) residues carboxyl (-) residues BD PureCoatTM Advantages : Chemically defined Animal-free igh consistency Low CV s in cell-based assays Stable at room temperature (3 years) or Thin-film coating Carboxyl or Amine momomers 21

22 Synthetic defined - Purecoat 22

23 Synthetic defined Purecoat ECM Mimetic 2 options : Fibronectine Collagen ECM Mimetic Surface technology Main applications : uman Mesenchymal Stem Cells uman Endothelial Progenitor Cells uman Keratinocytes 23

24 Synthetic defined PuraMatrix PuraMatrix Petpide ydrogel : - Synthetic matrix to create a defined 3D microenvironments origin: synthetic, animal free major components: - 16 aa peptide average pore size nm no growth factors - Applications : - Puramatrix scaffold allows creation of highly defined environment through addition of factors & compounds Does not activate RGD-dependent integrin signaling ighly reproducible experiments Compatible with most cells types in vitro & in vivo (stem cell proliferation, tumor cell migration and invasion, angiogenesis assays, tissue regeneration) 24

25 Synthetic Specific Synthemax Synthemax Surface : 1 Application Stem cell What is the Corning Synthemax Surface? polystyrene 1. Uncoated surface polystyrene 2. Acrylate coated polystyrene 3. Peptide covalently bound 25

26 Synthetic Specific Synthemax Advantages : Time Saving & Shelf Life Feeders Storage Shelf Life Easy-to-Use ECMs Synthemax Need to be used within a week Need to be stored at 70C When working they need to be kept cold No special storage conditions; can be stored at room temp Last only a week once prepared Plates need to be used as soon as possible 2 year shelf life arvest mouse embryos Grow up the Fibroblasts Time intensive Laborious timeconsuming prep No prep, just cut open the package and use 26

27 Synthetic Specific Synthemax Customer Needs Research Therapeutic Traceability 10-6 Sterility Consistent Performance Screening Shelf Life No Special Storage Conditions Ease of Use Purity Post-arvesting Lot Size Multiple Formats Inventory Availability Xeno-Free Substrate Ability to visualize cells Automation Friendly Format GMP Production Expand & Differentiate on same Surface 27

28 Synthetic Specific Synthemax Synthemax Consistency Defined, Synthetic, Xeno-Free Manufactured under GMP conditions Performs as good as Matrigel Consistent performance 2 year shelf life Defined Synthetic No animal components No prep, just cut open the package and use Easy-to-Use 28

29 Synthetic Specific Osteo Assay Osteo Assay Surface : 1 Application Osteo/bone assays Specifications : Inorganic synthetic coating that mimics in-vivo bone for in vitro bone cell assays Surface applications : - Consistent assay results for bone cell - Differentiation and function - Co-culture of bone cells / metastatic cancer cells - Alternative to preparing dentine, bone slices SEM image magnified not true representation 29

30 Synthetic Specific Osteo Assay Key Benefits Critical Attributes Differentiating Attributes Time Reduce time and cost of surface preparation Precoated, sterile, ready to use Direct Readout Detect both cell differentiation and functional analysis of bone cells on the same surface Supernatant can be sampled for cell function Transparent to conventional imaging techniques Amenable to automation Standard SBS format Can be used with standard imaging technologies Reliable cell response Consistent assays Standard process for manufacturing Uniform surface 30

31 Cell Culture surfaces Procuts available You can find the different cell culture surfaces on multiple vessels : Flasks Microplates from 6 to 1536 wells Dishes For specific applications : Assays microplates : Drug discovery Molecules screening Cells Culture inserts (Transwell ) : Coculture Migration Invasion 31

32 TOOLS TO FIND TE RIGT SURFACE Cell Culture Selection Guide Corning Cell Culture Selection Guide Corning Falcon Success Stories CellBIND Bibliography OsteoAssays Bibliography ULA (Ultra Low Attachment) Different Protocols or Application notes European Scientific Support (2 scientists) 32

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34 Glass Bottom Microplates Glass igh optical quality and scratch-resistant glass Bottom thickness 200 µm Intra well bottom flatness <50 µm Copolimer Cyclic Olefin Copolymer (COC) Bottom thickness: 127 µm Intra-well bottom flatness 5 µm Inter-well bottom flatness 30 µm 34

35 Flowell 35

36 Flowell 36

37 Axygen 5 ml Snaplock Microtubes 5mL 15mm No necesita adaptador Rotores 15 mm (15ml) Rotores 17 mm (15ml) rpm 120ºC (15 15psi) Libre de ARNasas, ADNasas 37

38 Corning Life Science - AXYGEN Biosciences Thanks a lot for your time and attention. QUESTIONS? 38