Transwell Permeable Supports - Corning

1 2007 Corning Incorporated Printed in USA9/07 CLS-CC-011 REV4 Corning and Transwell are registered trademarks of Corning , Incorporated, Corning , NY. Netwell and Snapwell are trademarks of Corning , Incorporated, Corning , NY. Corning Incorporated, One Riverfront Plaza, Corning , NY 14831-0001 Corning IncorporatedLife SciencesTower 2, 4th Floor900 Chelmsford , MA 01851t Support OfficesASIA/PACIFICA ustraliat 61 2-9416-0492f 61 2-9416-0493 Chinat 86 21-3222-4666f 86 21-6288-1575 Hong Kongt 852-2807-2723f 852-2807-2152 Indiat 91-124-235 7850f 91-124-401 0207 Japant 81 (0) 3-3586 1996/1997f 81 (0) 3-3586 1291/1292 Koreat 82 2-796-9500f 82 2-796-9300 Singaporet 65 6733-6511f 65 6861-2913 Taiwant 886 2-2716-0338f 886 2-2716-0339 EUROPEF rancet 0800 916 882f 0800 918 636 Germanyt 0800 101 1153f 0800 101 2427 The Netherlandst 31 20 655 79 28f 31 20 659 76 73 United Kingdomt 0800 376 8660f 0800 279 1117 All Other European Countriest 31 (0) 20 659 60 51f 31 (0) 20 659 76 73 LATIN AMERICAB rasilt (55-11) 3089-7419f (55-11) 3167-0700 Mexicot (52-81)

2 8158-8400f (52-81) 8313-8589 Transwell Permeable SupportsIncluding Snapwell and Netwell InsertsInstructions for Use1 INTRODUCTIONT ranswell Permeable Supports are convenient and easy-to-usedevices for studies of both anchorage-dependent and anchorage-independent cells. These inserts provide independent access toboth sides of a monolayer, thus giving researchers a versatile toolto study transport and other metabolic activities in vitro. Transwellunits are available in a range of diameters, membrane types andpore sizes to satisfy a variety of research INSERT PRODUCT DESCRIPTIONSS tandard Transwell InsertsStandard Transwell inserts are available in four membranediameters: mm (24 well plate), 12 mm (12 well plate), 24 mm(6 well plate) and 75 mm (100 mm dish) formats. See Table 1 forcell growth areas. Table 1. Transwell Permeable Support Growth Areas* Transwell Multiple Well Plate Insert Membrane Insert Diameter or Dish StyleGrowth Area mm24 cm212 mm12 cm224 mm6 cm275 mm100 mm dish44 cm2*Values are reported as nominal and may vary slightly due to inherent variabilityof our manufacturing process.

3 To insure success, we recommend that end usersvalidate their methods independent from our reported OF CONTENTSI ntroduction .. 1 Transwell Insert Product Descriptions .. 1 Standard Transwell Inserts .. 1 Membrane Types .. 2 Pore Sizes .. 3 Pore Density .. 4 Chemical Compatibility .. 4 Using Transwell Permeable Supports .. 5 Directions for Use .. 5 Helpful Hints .. 6 Additional Permeable Support Products .. 7 HTS Transwell -24 Well Permeable Supports .. 7 HTS Transwell -96 Well Permeable Supports .. 8 Snapwell Inserts .. 8 Netwell Inserts .. 9 Technical Assistance .. 9 Ordering Information .. 10 Polycarbonate Transwell Inserts .. 10 Polyester Transwell Inserts .. 10 collagen Coated Transwell -COL Inserts .. 11 HTS Transwell 24 System .. 11 HTS Transwell -96 System .. 12 Snapwell Inserts .. 12 Netwell Inserts .. 136, 12, and 24 Well Cell culture Plates.

4 1332 Polycarbonate Transwell inserts feature thin, translucent mem-branes available in six pore sizes ranging from m to are treated for optimal cell attachment. They are suppliedsterile and come preloaded in multiple well plates or dishes. The polycarbonate membrane is compatible with most organicfixatives and stains. Polyester (PET) Transwell -Clear inserts have microscopicallytransparent polyester membranes that are tissue culture treatedfor optimal cell attachment and growth. Transwell -Clear insertsprovide better cell visibility under phase contrast microscopy andallow assessment of cell viability and monolayer formation. Transwell -COL inserts have transparent (when wet), collagen -treated PTFE membranes that promote cell attachment andspreading and allow cells to be visualized during culture . TheTranswell-COL membranes are coated with an equimolar mix-ture of types I and III collagen derived from bovine traditional coating techniques that result in occluding film layers, Corning s proprietary coating process results in abiologically stable collagen that covers every fibril of the filtermatrix, thereby retaining the porosity of the SizesSelecting the correct pore size for experiments is also very impor-tant.

5 The smaller pore size Transwell membranes ( or m)are primarily used in drug transport studies. Cell invasion, chemo-taxis and motility studies are usually done in Transwell membraneswith m or larger pores. The ability of cells to migratethrough pores of a membrane is dependent on the cell line usedand the culture conditions, as well as the pore size. Cell migra-tion will not occur with pores smaller than m. For criticalexperiments, we suggest testing with appropriate controls and arange of pores sizes to determine which size works best with yourcell cultures and your specific application. As an alternative, fol-low recommendations from the scientific literature. For additionalapplication and use information, please refer to the TranswellSeveral membranetypes and a largeselection of poresizes are availablewith each of theseunits.

6 A patented*self-centeringdesign preventsmedium fromwicking between the sides of the insert and the well wall. Thehanging design keeps the Transwell membrane about a milli-meter off the bottom of the well. This prevents co-cultured cellmonolayers in the bottom of the well from being scratched ordisturbed when the insert is moved. Windows or openings in thesides of the Transwell insert allow access to the lower TypesTranswell Permeable Supports are available in three membranematerials: polycarbonate, polyester (PET), and collagen -coatedpolytetrafluoroethylene (PTFE). See Table 2 for additionalinformation on these membrane 2. Characteristics of Transwell MembranesPolyester PTFE/Characteristics(PET)PolycarbonateCo llagen-coatedOptical propertiesClear Translucent Clear when wetCell visibilityGoodPoorCell outlinesTissue culture treatedYesYesNoNominal Membrane 10 m10 m30 mthickness Matrix/ECM coatableYesYesYesCollagen treatedNoNoYesAvailable Pore Sizes ( m) , , , , , , , Membrane manufacturing processes are inherently variable; thus, actual values may vary up to 60% of nominal depending on membrane type and measurement techniques.

7 *Patent number 5,026, membraneLower compartmentTranswell insertUpper compartment54 USING Transwell Permeable SUPPORTSD irections for Use1. Transwell inserts are used by first adding medium to the multiplewell plate well, followed by adding the Transwell inserts, andlastly adding the medium and cells to the inside medium volumes are shown in Table 4. Recommended Transwell Permeable Support Medium VolumesTranswell Insert Multiple Well Volume Volume Added Insert Membrane Plate or Added per to Inside of DiameterGrowth AreaDish TypePlate WellTranswell mm cm296 mL mm cm224 mL12 mm cm212 mL24 mm cm26 mL75 mm44 cm2100 mm dish 13 mL2. An initial equilibrium period may be used to improve cellattachment by adding medium to the multiple well plate welland then to the Transwell insert. The plate should then beincubated for at least one hour or even overnight at the sametemperature that will be used to grow the cells.

8 The cells arethen added in fresh medium to the Transwell insert andreturned to the The medium level should be checked periodically and freshmedium added as Transwell inserts have three openings for standard pipette tipsto allow samples to be added or removed from the Cell monolayers may be fixed and stained while in the Transwellinsert using standard cytological techniques. Avoid using sol-vents that dissolve polystyrene or polycarbonate or polyestermembrane materials. Processing steps may be carried out byinserts bibliography on the Technical Information section of theCorning Life Sciences web site ( lifesciences)that lists over 800 literature references using Transwell permeablesupports. Pore DensityOf the three types of Transwell membranes, only the PTFE membrane does not have a defined pore density because it is atortuous path membrane.

9 The two membranes with nominallydefined pore densities are polycarbonate and polyester. The poly-ester Transwell membranes do not have as high a pore density asthe polycarbonate Transwell membranes but they have betteroptical clarity as a result. The nominal pore densities for poly-carbonate and polyester membranes are given in Table 3. Nominal Pore Densities for Transwell Polyesterand Polycarbonate MembranesNominal Pore DensityPore Size Polycarbonate Membrane Polyester m1 x 108pores/cm24 x 106 m x 106 m2 x 106pores/cm22 x 106 m4 x 105pores/cm2 m1 x 105pores/cm21 x 105pores/cm2 Chemical CompatibilityAll of the Transwell membranes are compatible with histologicalfixatives including methanol and formaldehyde. The polyesterTranswell membranes have the best overall chemical membranes (but not the polystyrene housings) are com-patible with many alcohols, amines, esters, ethers, ketones, oilsand some solvents including many halogenated hydrocarbons andDMSO, but are not recommended for use with strong acids Cells requiring extracellular matrix coatings on plasticsubstrates will also require them on Permeable The Transwell -Clear insert contains a transparent tissueculture treated polyester membrane that allows easy viewing of cells using phase contrast microscopy.

10 7. The Transwell -COL insert contains a PTFE membrane thathas been treated with an equimolar mixture of types I and IIIbovine placental collagens. This results in a biologicallystabilized collagen matrix covering the fibrils of the filtermembrane. These Transwell inserts are excellent for thegrowth of cells requiring a biological Permeable SUPPORTPRODUCTSHTS Transwell -24 SystemThe HTS Transwell -24 System is an array of 24 indi-vidual Transwell inserts ( diameter membranes)connected by a rigid, robotics-friendly holder that enablesall 24 Transwell inserts to behandled as a single unit. Thismakes the HTS Transwell -24 System an ideal tool for running automated, high throughputdrug transport (Caco-2 cells) or cell toxicity studies. The tissueculture treated polycarbonate membrane is available in either m or m pore sizes. The tissue culture treated polyestermembrane is available in m pore size.