Lysis:Article Title: Pyrrolopyrimidine and pyrrolopyridine derivatives
Article Snippet: .. 10 mM (Sigma Aldrich; Cat. No. E-4884) RPMI 1640 (Lonza Cat. No. BE12-115F/U1) FBS (Sigma Cat. No. F7524, heat inactivated 30′/56° C.) Dimethyl sulfoxide (DMSO) (Sigma, Cat. No. D2650) Cell lysis buffer (pH 7.4): PBS+1% Triton X-100 (Sigma Aldrich; Cat. No. X100)—10 ml Phospho-Stop—1 tablet (Sigma, Cat. No. 4693124001) Protease Inhibitor—1 tablet (Sigma, Cat. No. 4906837001) Hydrogen peroxide solution 3% w/w (Sigma Aldrich, Cat. No. H-6520) PBS (Sigma Aldrich, Cat. No. P4417-100TAB) Microplate, 96 well, pp, v-bottom, clear (Greiner bio-one; Cat. No. 651201) Tissue culture plate, 6 well, flat bottom with low evaporation lid (Falcon, Cat. No. 353224) 50 mL falcon tube (TPP, Cat. No. 91050) Pierce BCA Protein Assay Kit (Thermo Scientific, Cat. No. 23225) Immulon® MicrotiterTM 96-Well Plate 1B (Thermo Scientific, Cat. No. 3355) Wes 12-230 kDa Master Kit (Protein Simple, Cat. No. PS-MK14) MKK3 (D4C3) Rabbit mAb (Cell Signaling, Cat. No. 8535) Phospho-MKK3 (Ser189)/MKK6 (Ser207) (D8E9) Rabbit mAb (Cell Signaling, Cat. No. 12280) The assay was configured to run using the following conditions: Tested compounds concentration range: 1-0.0014 μM (7 three-fold serial dilutions); H2O2 concentration: 3 mM ..
Evaporation:Article Title: Pyrrolopyrimidine and pyrrolopyridine derivatives
Article Snippet: .. 10 mM (Sigma Aldrich; Cat. No. E-4884) RPMI 1640 (Lonza Cat. No. BE12-115F/U1) FBS (Sigma Cat. No. F7524, heat inactivated 30′/56° C.) Dimethyl sulfoxide (DMSO) (Sigma, Cat. No. D2650) Cell lysis buffer (pH 7.4): PBS+1% Triton X-100 (Sigma Aldrich; Cat. No. X100)—10 ml Phospho-Stop—1 tablet (Sigma, Cat. No. 4693124001) Protease Inhibitor—1 tablet (Sigma, Cat. No. 4906837001) Hydrogen peroxide solution 3% w/w (Sigma Aldrich, Cat. No. H-6520) PBS (Sigma Aldrich, Cat. No. P4417-100TAB) Microplate, 96 well, pp, v-bottom, clear (Greiner bio-one; Cat. No. 651201) Tissue culture plate, 6 well, flat bottom with low evaporation lid (Falcon, Cat. No. 353224) 50 mL falcon tube (TPP, Cat. No. 91050) Pierce BCA Protein Assay Kit (Thermo Scientific, Cat. No. 23225) Immulon® MicrotiterTM 96-Well Plate 1B (Thermo Scientific, Cat. No. 3355) Wes 12-230 kDa Master Kit (Protein Simple, Cat. No. PS-MK14) MKK3 (D4C3) Rabbit mAb (Cell Signaling, Cat. No. 8535) Phospho-MKK3 (Ser189)/MKK6 (Ser207) (D8E9) Rabbit mAb (Cell Signaling, Cat. No. 12280) The assay was configured to run using the following conditions: Tested compounds concentration range: 1-0.0014 μM (7 three-fold serial dilutions); H2O2 concentration: 3 mM ..
Bicinchoninic Acid Protein Assay:Article Title: Pyrrolopyrimidine and pyrrolopyridine derivatives
Article Snippet: .. 10 mM (Sigma Aldrich; Cat. No. E-4884) RPMI 1640 (Lonza Cat. No. BE12-115F/U1) FBS (Sigma Cat. No. F7524, heat inactivated 30′/56° C.) Dimethyl sulfoxide (DMSO) (Sigma, Cat. No. D2650) Cell lysis buffer (pH 7.4): PBS+1% Triton X-100 (Sigma Aldrich; Cat. No. X100)—10 ml Phospho-Stop—1 tablet (Sigma, Cat. No. 4693124001) Protease Inhibitor—1 tablet (Sigma, Cat. No. 4906837001) Hydrogen peroxide solution 3% w/w (Sigma Aldrich, Cat. No. H-6520) PBS (Sigma Aldrich, Cat. No. P4417-100TAB) Microplate, 96 well, pp, v-bottom, clear (Greiner bio-one; Cat. No. 651201) Tissue culture plate, 6 well, flat bottom with low evaporation lid (Falcon, Cat. No. 353224) 50 mL falcon tube (TPP, Cat. No. 91050) Pierce BCA Protein Assay Kit (Thermo Scientific, Cat. No. 23225) Immulon® MicrotiterTM 96-Well Plate 1B (Thermo Scientific, Cat. No. 3355) Wes 12-230 kDa Master Kit (Protein Simple, Cat. No. PS-MK14) MKK3 (D4C3) Rabbit mAb (Cell Signaling, Cat. No. 8535) Phospho-MKK3 (Ser189)/MKK6 (Ser207) (D8E9) Rabbit mAb (Cell Signaling, Cat. No. 12280) The assay was configured to run using the following conditions: Tested compounds concentration range: 1-0.0014 μM (7 three-fold serial dilutions); H2O2 concentration: 3 mM ..
Concentration Assay:Article Title: Pyrrolopyrimidine and pyrrolopyridine derivatives
Article Snippet: .. 10 mM (Sigma Aldrich; Cat. No. E-4884) RPMI 1640 (Lonza Cat. No. BE12-115F/U1) FBS (Sigma Cat. No. F7524, heat inactivated 30′/56° C.) Dimethyl sulfoxide (DMSO) (Sigma, Cat. No. D2650) Cell lysis buffer (pH 7.4): PBS+1% Triton X-100 (Sigma Aldrich; Cat. No. X100)—10 ml Phospho-Stop—1 tablet (Sigma, Cat. No. 4693124001) Protease Inhibitor—1 tablet (Sigma, Cat. No. 4906837001) Hydrogen peroxide solution 3% w/w (Sigma Aldrich, Cat. No. H-6520) PBS (Sigma Aldrich, Cat. No. P4417-100TAB) Microplate, 96 well, pp, v-bottom, clear (Greiner bio-one; Cat. No. 651201) Tissue culture plate, 6 well, flat bottom with low evaporation lid (Falcon, Cat. No. 353224) 50 mL falcon tube (TPP, Cat. No. 91050) Pierce BCA Protein Assay Kit (Thermo Scientific, Cat. No. 23225) Immulon® MicrotiterTM 96-Well Plate 1B (Thermo Scientific, Cat. No. 3355) Wes 12-230 kDa Master Kit (Protein Simple, Cat. No. PS-MK14) MKK3 (D4C3) Rabbit mAb (Cell Signaling, Cat. No. 8535) Phospho-MKK3 (Ser189)/MKK6 (Ser207) (D8E9) Rabbit mAb (Cell Signaling, Cat. No. 12280) The assay was configured to run using the following conditions: Tested compounds concentration range: 1-0.0014 μM (7 three-fold serial dilutions); H2O2 concentration: 3 mM ..
Transferring:Article Title: FRET Reporter Assays for cAMP and Calcium in a 96-well Format Using Genetically Encoded Biosensors Expressed in Living Cells
Article Snippet: .. 10 ml Serological Pipet (Krackeler, catalog number: 229010B) 5 ml Serological Pipet (Krackeler, catalog number: 229005B) 1.5 ml Centrifuge Tube (Axygen, catalog number: MCT-150-C) 15 ml Centrifuge Tube (Krackeler, catalog number: 229411) 50 ml Centrifuge Tube (Krackeler, catalog number: 229421) 100 mm x 20 mm Tissue Culture Dish (Krackeler, catalog number: 229621) 96-Well Plate Black, Clear Bottom (Corning, catalog number: 3603) 96-Well Plate, V-Bottom (Greiner Bio-One, catalog number: 651201) FlexStation Pipette Tips (Black, 96) (Molecular Devices, catalog number: 9000–0911) Finntip Flex 300 μl Sterile Pipette Tips (Thermo Scientific, catalog number: 94060513) Channelmate 25 ml Inserts (USA Scientific, catalog number: 1346–2510) Multiwell Plate Washer/dispenser Manifold (Sigma, catalog number: M2781–1EA) 0.2 μm PES Membrane Filter for Vacuum Filtration HEK293/hNPY2R/H188 c18 cells expressing human NPY2R receptor and H188 HEK293/hGCGR/H188 c10 cells expressing human glucagon receptor and H188 HEK293/hCCK 2 R c29 cells expressing human CCK 2 R only Sodium Chloride (Fisher Scientific, catalog number: BP358–1) Sodium Hydroxide (Fisher Scientific, catalog number: BP359–500) Potassium Chloride (Fisher Scientific, catalog number: BP366–500) Magnesium Chloride Hexahydrate (Fisher Scientific, catalog number: BP214–500) Calcium Chloride Dihydrate (Fisher Scientific, catalog number: BP510–100) D-Glucose (Sigma, catalog number: G5767–500G) HEPES 1 M Solution (Sigma, catalog number: H0887; store at 4 °C) Bovine Serum Albumin (Sigma, catalog number: A7030–100G; store at 4 °C) Rat Tail Collagen (Life Technologies, catalog number: A10483–01; store at 4 °C) Glacial Acetic Acid (Thermo Scientific, catalog number: A38S-500) Phosphate-Buffered Saline, 1x (Corning, catalog number: 21–040-CV) DMSO (Sigma, catalog number: D8418–100ML) Fetal Bovine Serum (Sigma, catalog number: 12303C-500ML; store at −20 °C) Dulbecco’s Modified Eagle Medium (Sigma, catalog number: D6429–500ML; store at 4 °C) Trypsin-EDTA (0.25%) (Thermo Scientific, catalog number: 25200072; store at −20 °C) Pen Strep (Life Technologies, catalog number: 15140–122; store at −20 °C) G-418 Disulfate Salt Solution (Sigma, catalog number: G8168–10ML; store at 4 °C) Forskolin (Sigma, catalog number: F6886–10MG) Glucagon (Sigma, catalog number: G1774-.1MG; store at −20 °C) Demogastrin-2 (DG2) (Pichem, Bat. .. No. M24–061202; store at −20 °C) PYY(3–36) (Sigma, catalog number: P220–500UG; store at −20 °C) Standard Extracellular Saline (SES) Solution, pH 7.4 (see Recipes ) Rat Tail Collagen (RTC) Coating Solution (for one 96-well plate) (see Recipes )
Sterility:Article Title: FRET Reporter Assays for cAMP and Calcium in a 96-well Format Using Genetically Encoded Biosensors Expressed in Living Cells
Article Snippet: .. 10 ml Serological Pipet (Krackeler, catalog number: 229010B) 5 ml Serological Pipet (Krackeler, catalog number: 229005B) 1.5 ml Centrifuge Tube (Axygen, catalog number: MCT-150-C) 15 ml Centrifuge Tube (Krackeler, catalog number: 229411) 50 ml Centrifuge Tube (Krackeler, catalog number: 229421) 100 mm x 20 mm Tissue Culture Dish (Krackeler, catalog number: 229621) 96-Well Plate Black, Clear Bottom (Corning, catalog number: 3603) 96-Well Plate, V-Bottom (Greiner Bio-One, catalog number: 651201) FlexStation Pipette Tips (Black, 96) (Molecular Devices, catalog number: 9000–0911) Finntip Flex 300 μl Sterile Pipette Tips (Thermo Scientific, catalog number: 94060513) Channelmate 25 ml Inserts (USA Scientific, catalog number: 1346–2510) Multiwell Plate Washer/dispenser Manifold (Sigma, catalog number: M2781–1EA) 0.2 μm PES Membrane Filter for Vacuum Filtration HEK293/hNPY2R/H188 c18 cells expressing human NPY2R receptor and H188 HEK293/hGCGR/H188 c10 cells expressing human glucagon receptor and H188 HEK293/hCCK 2 R c29 cells expressing human CCK 2 R only Sodium Chloride (Fisher Scientific, catalog number: BP358–1) Sodium Hydroxide (Fisher Scientific, catalog number: BP359–500) Potassium Chloride (Fisher Scientific, catalog number: BP366–500) Magnesium Chloride Hexahydrate (Fisher Scientific, catalog number: BP214–500) Calcium Chloride Dihydrate (Fisher Scientific, catalog number: BP510–100) D-Glucose (Sigma, catalog number: G5767–500G) HEPES 1 M Solution (Sigma, catalog number: H0887; store at 4 °C) Bovine Serum Albumin (Sigma, catalog number: A7030–100G; store at 4 °C) Rat Tail Collagen (Life Technologies, catalog number: A10483–01; store at 4 °C) Glacial Acetic Acid (Thermo Scientific, catalog number: A38S-500) Phosphate-Buffered Saline, 1x (Corning, catalog number: 21–040-CV) DMSO (Sigma, catalog number: D8418–100ML) Fetal Bovine Serum (Sigma, catalog number: 12303C-500ML; store at −20 °C) Dulbecco’s Modified Eagle Medium (Sigma, catalog number: D6429–500ML; store at 4 °C) Trypsin-EDTA (0.25%) (Thermo Scientific, catalog number: 25200072; store at −20 °C) Pen Strep (Life Technologies, catalog number: 15140–122; store at −20 °C) G-418 Disulfate Salt Solution (Sigma, catalog number: G8168–10ML; store at 4 °C) Forskolin (Sigma, catalog number: F6886–10MG) Glucagon (Sigma, catalog number: G1774-.1MG; store at −20 °C) Demogastrin-2 (DG2) (Pichem, Bat. .. No. M24–061202; store at −20 °C) PYY(3–36) (Sigma, catalog number: P220–500UG; store at −20 °C) Standard Extracellular Saline (SES) Solution, pH 7.4 (see Recipes ) Rat Tail Collagen (RTC) Coating Solution (for one 96-well plate) (see Recipes )
Article Title: Use of standard U-bottom and V-bottom well plates to generate neuroepithelial embryoid bodies
Article Snippet: .. A 100 μL volume of anti-adherence rinsing solution (StemCell Technologies, cat. no. 07010; Canada) was added to untreated sterile CellStar U-bottom or V-bottom ( ) 96-well plates (Greiner Bio-One, cat. no. 650185(U), 651161(V); Austria). .. After incubation for 5 min at room temperature, the anti-adherence rinsing solution was aspirated, and the wells were washed with Dulbecco’s phosphate buffered saline (DPBS) (Gibco, cat. no. 14190250; Carlsbad, CA, USA) for another 5 min at room temperature.
Article Title: Use of standard U-bottom and V-bottom well plates to generate neuroepithelial embryoid bodies
Article Snippet: .. A 100 μL volume of anti-adherence rinsing solution (StemCell Technologies, Vancouver, Canada, cat. no. 07010) was added to untreated sterile CellStar U-bottom or V-bottom ( ) 96-well plates (Greiner Bio-One, Kremsmuenster, Austria, cat. no. 650185(U), 651161(V)). .. After incubation for 5 min at room temperature, the anti-adherence rinsing solution was aspirated, and the wells were washed with Dulbecco’s phosphate buffered saline (DPBS) (Gibco, Carlsbad, CA, United States, cat. no. 14190250) for another 5 min at room temperature.
Membrane:Article Title: FRET Reporter Assays for cAMP and Calcium in a 96-well Format Using Genetically Encoded Biosensors Expressed in Living Cells
Article Snippet: .. 10 ml Serological Pipet (Krackeler, catalog number: 229010B) 5 ml Serological Pipet (Krackeler, catalog number: 229005B) 1.5 ml Centrifuge Tube (Axygen, catalog number: MCT-150-C) 15 ml Centrifuge Tube (Krackeler, catalog number: 229411) 50 ml Centrifuge Tube (Krackeler, catalog number: 229421) 100 mm x 20 mm Tissue Culture Dish (Krackeler, catalog number: 229621) 96-Well Plate Black, Clear Bottom (Corning, catalog number: 3603) 96-Well Plate, V-Bottom (Greiner Bio-One, catalog number: 651201) FlexStation Pipette Tips (Black, 96) (Molecular Devices, catalog number: 9000–0911) Finntip Flex 300 μl Sterile Pipette Tips (Thermo Scientific, catalog number: 94060513) Channelmate 25 ml Inserts (USA Scientific, catalog number: 1346–2510) Multiwell Plate Washer/dispenser Manifold (Sigma, catalog number: M2781–1EA) 0.2 μm PES Membrane Filter for Vacuum Filtration HEK293/hNPY2R/H188 c18 cells expressing human NPY2R receptor and H188 HEK293/hGCGR/H188 c10 cells expressing human glucagon receptor and H188 HEK293/hCCK 2 R c29 cells expressing human CCK 2 R only Sodium Chloride (Fisher Scientific, catalog number: BP358–1) Sodium Hydroxide (Fisher Scientific, catalog number: BP359–500) Potassium Chloride (Fisher Scientific, catalog number: BP366–500) Magnesium Chloride Hexahydrate (Fisher Scientific, catalog number: BP214–500) Calcium Chloride Dihydrate (Fisher Scientific, catalog number: BP510–100) D-Glucose (Sigma, catalog number: G5767–500G) HEPES 1 M Solution (Sigma, catalog number: H0887; store at 4 °C) Bovine Serum Albumin (Sigma, catalog number: A7030–100G; store at 4 °C) Rat Tail Collagen (Life Technologies, catalog number: A10483–01; store at 4 °C) Glacial Acetic Acid (Thermo Scientific, catalog number: A38S-500) Phosphate-Buffered Saline, 1x (Corning, catalog number: 21–040-CV) DMSO (Sigma, catalog number: D8418–100ML) Fetal Bovine Serum (Sigma, catalog number: 12303C-500ML; store at −20 °C) Dulbecco’s Modified Eagle Medium (Sigma, catalog number: D6429–500ML; store at 4 °C) Trypsin-EDTA (0.25%) (Thermo Scientific, catalog number: 25200072; store at −20 °C) Pen Strep (Life Technologies, catalog number: 15140–122; store at −20 °C) G-418 Disulfate Salt Solution (Sigma, catalog number: G8168–10ML; store at 4 °C) Forskolin (Sigma, catalog number: F6886–10MG) Glucagon (Sigma, catalog number: G1774-.1MG; store at −20 °C) Demogastrin-2 (DG2) (Pichem, Bat. .. No. M24–061202; store at −20 °C) PYY(3–36) (Sigma, catalog number: P220–500UG; store at −20 °C) Standard Extracellular Saline (SES) Solution, pH 7.4 (see Recipes ) Rat Tail Collagen (RTC) Coating Solution (for one 96-well plate) (see Recipes )
Suction Filtration:Article Title: FRET Reporter Assays for cAMP and Calcium in a 96-well Format Using Genetically Encoded Biosensors Expressed in Living Cells
Article Snippet: .. 10 ml Serological Pipet (Krackeler, catalog number: 229010B) 5 ml Serological Pipet (Krackeler, catalog number: 229005B) 1.5 ml Centrifuge Tube (Axygen, catalog number: MCT-150-C) 15 ml Centrifuge Tube (Krackeler, catalog number: 229411) 50 ml Centrifuge Tube (Krackeler, catalog number: 229421) 100 mm x 20 mm Tissue Culture Dish (Krackeler, catalog number: 229621) 96-Well Plate Black, Clear Bottom (Corning, catalog number: 3603) 96-Well Plate, V-Bottom (Greiner Bio-One, catalog number: 651201) FlexStation Pipette Tips (Black, 96) (Molecular Devices, catalog number: 9000–0911) Finntip Flex 300 μl Sterile Pipette Tips (Thermo Scientific, catalog number: 94060513) Channelmate 25 ml Inserts (USA Scientific, catalog number: 1346–2510) Multiwell Plate Washer/dispenser Manifold (Sigma, catalog number: M2781–1EA) 0.2 μm PES Membrane Filter for Vacuum Filtration HEK293/hNPY2R/H188 c18 cells expressing human NPY2R receptor and H188 HEK293/hGCGR/H188 c10 cells expressing human glucagon receptor and H188 HEK293/hCCK 2 R c29 cells expressing human CCK 2 R only Sodium Chloride (Fisher Scientific, catalog number: BP358–1) Sodium Hydroxide (Fisher Scientific, catalog number: BP359–500) Potassium Chloride (Fisher Scientific, catalog number: BP366–500) Magnesium Chloride Hexahydrate (Fisher Scientific, catalog number: BP214–500) Calcium Chloride Dihydrate (Fisher Scientific, catalog number: BP510–100) D-Glucose (Sigma, catalog number: G5767–500G) HEPES 1 M Solution (Sigma, catalog number: H0887; store at 4 °C) Bovine Serum Albumin (Sigma, catalog number: A7030–100G; store at 4 °C) Rat Tail Collagen (Life Technologies, catalog number: A10483–01; store at 4 °C) Glacial Acetic Acid (Thermo Scientific, catalog number: A38S-500) Phosphate-Buffered Saline, 1x (Corning, catalog number: 21–040-CV) DMSO (Sigma, catalog number: D8418–100ML) Fetal Bovine Serum (Sigma, catalog number: 12303C-500ML; store at −20 °C) Dulbecco’s Modified Eagle Medium (Sigma, catalog number: D6429–500ML; store at 4 °C) Trypsin-EDTA (0.25%) (Thermo Scientific, catalog number: 25200072; store at −20 °C) Pen Strep (Life Technologies, catalog number: 15140–122; store at −20 °C) G-418 Disulfate Salt Solution (Sigma, catalog number: G8168–10ML; store at 4 °C) Forskolin (Sigma, catalog number: F6886–10MG) Glucagon (Sigma, catalog number: G1774-.1MG; store at −20 °C) Demogastrin-2 (DG2) (Pichem, Bat. .. No. M24–061202; store at −20 °C) PYY(3–36) (Sigma, catalog number: P220–500UG; store at −20 °C) Standard Extracellular Saline (SES) Solution, pH 7.4 (see Recipes ) Rat Tail Collagen (RTC) Coating Solution (for one 96-well plate) (see Recipes )
Expressing:Article Title: FRET Reporter Assays for cAMP and Calcium in a 96-well Format Using Genetically Encoded Biosensors Expressed in Living Cells
Article Snippet: .. 10 ml Serological Pipet (Krackeler, catalog number: 229010B) 5 ml Serological Pipet (Krackeler, catalog number: 229005B) 1.5 ml Centrifuge Tube (Axygen, catalog number: MCT-150-C) 15 ml Centrifuge Tube (Krackeler, catalog number: 229411) 50 ml Centrifuge Tube (Krackeler, catalog number: 229421) 100 mm x 20 mm Tissue Culture Dish (Krackeler, catalog number: 229621) 96-Well Plate Black, Clear Bottom (Corning, catalog number: 3603) 96-Well Plate, V-Bottom (Greiner Bio-One, catalog number: 651201) FlexStation Pipette Tips (Black, 96) (Molecular Devices, catalog number: 9000–0911) Finntip Flex 300 μl Sterile Pipette Tips (Thermo Scientific, catalog number: 94060513) Channelmate 25 ml Inserts (USA Scientific, catalog number: 1346–2510) Multiwell Plate Washer/dispenser Manifold (Sigma, catalog number: M2781–1EA) 0.2 μm PES Membrane Filter for Vacuum Filtration HEK293/hNPY2R/H188 c18 cells expressing human NPY2R receptor and H188 HEK293/hGCGR/H188 c10 cells expressing human glucagon receptor and H188 HEK293/hCCK 2 R c29 cells expressing human CCK 2 R only Sodium Chloride (Fisher Scientific, catalog number: BP358–1) Sodium Hydroxide (Fisher Scientific, catalog number: BP359–500) Potassium Chloride (Fisher Scientific, catalog number: BP366–500) Magnesium Chloride Hexahydrate (Fisher Scientific, catalog number: BP214–500) Calcium Chloride Dihydrate (Fisher Scientific, catalog number: BP510–100) D-Glucose (Sigma, catalog number: G5767–500G) HEPES 1 M Solution (Sigma, catalog number: H0887; store at 4 °C) Bovine Serum Albumin (Sigma, catalog number: A7030–100G; store at 4 °C) Rat Tail Collagen (Life Technologies, catalog number: A10483–01; store at 4 °C) Glacial Acetic Acid (Thermo Scientific, catalog number: A38S-500) Phosphate-Buffered Saline, 1x (Corning, catalog number: 21–040-CV) DMSO (Sigma, catalog number: D8418–100ML) Fetal Bovine Serum (Sigma, catalog number: 12303C-500ML; store at −20 °C) Dulbecco’s Modified Eagle Medium (Sigma, catalog number: D6429–500ML; store at 4 °C) Trypsin-EDTA (0.25%) (Thermo Scientific, catalog number: 25200072; store at −20 °C) Pen Strep (Life Technologies, catalog number: 15140–122; store at −20 °C) G-418 Disulfate Salt Solution (Sigma, catalog number: G8168–10ML; store at 4 °C) Forskolin (Sigma, catalog number: F6886–10MG) Glucagon (Sigma, catalog number: G1774-.1MG; store at −20 °C) Demogastrin-2 (DG2) (Pichem, Bat. .. No. M24–061202; store at −20 °C) PYY(3–36) (Sigma, catalog number: P220–500UG; store at −20 °C) Standard Extracellular Saline (SES) Solution, pH 7.4 (see Recipes ) Rat Tail Collagen (RTC) Coating Solution (for one 96-well plate) (see Recipes )
Saline:Article Title: FRET Reporter Assays for cAMP and Calcium in a 96-well Format Using Genetically Encoded Biosensors Expressed in Living Cells
Article Snippet: .. 10 ml Serological Pipet (Krackeler, catalog number: 229010B) 5 ml Serological Pipet (Krackeler, catalog number: 229005B) 1.5 ml Centrifuge Tube (Axygen, catalog number: MCT-150-C) 15 ml Centrifuge Tube (Krackeler, catalog number: 229411) 50 ml Centrifuge Tube (Krackeler, catalog number: 229421) 100 mm x 20 mm Tissue Culture Dish (Krackeler, catalog number: 229621) 96-Well Plate Black, Clear Bottom (Corning, catalog number: 3603) 96-Well Plate, V-Bottom (Greiner Bio-One, catalog number: 651201) FlexStation Pipette Tips (Black, 96) (Molecular Devices, catalog number: 9000–0911) Finntip Flex 300 μl Sterile Pipette Tips (Thermo Scientific, catalog number: 94060513) Channelmate 25 ml Inserts (USA Scientific, catalog number: 1346–2510) Multiwell Plate Washer/dispenser Manifold (Sigma, catalog number: M2781–1EA) 0.2 μm PES Membrane Filter for Vacuum Filtration HEK293/hNPY2R/H188 c18 cells expressing human NPY2R receptor and H188 HEK293/hGCGR/H188 c10 cells expressing human glucagon receptor and H188 HEK293/hCCK 2 R c29 cells expressing human CCK 2 R only Sodium Chloride (Fisher Scientific, catalog number: BP358–1) Sodium Hydroxide (Fisher Scientific, catalog number: BP359–500) Potassium Chloride (Fisher Scientific, catalog number: BP366–500) Magnesium Chloride Hexahydrate (Fisher Scientific, catalog number: BP214–500) Calcium Chloride Dihydrate (Fisher Scientific, catalog number: BP510–100) D-Glucose (Sigma, catalog number: G5767–500G) HEPES 1 M Solution (Sigma, catalog number: H0887; store at 4 °C) Bovine Serum Albumin (Sigma, catalog number: A7030–100G; store at 4 °C) Rat Tail Collagen (Life Technologies, catalog number: A10483–01; store at 4 °C) Glacial Acetic Acid (Thermo Scientific, catalog number: A38S-500) Phosphate-Buffered Saline, 1x (Corning, catalog number: 21–040-CV) DMSO (Sigma, catalog number: D8418–100ML) Fetal Bovine Serum (Sigma, catalog number: 12303C-500ML; store at −20 °C) Dulbecco’s Modified Eagle Medium (Sigma, catalog number: D6429–500ML; store at 4 °C) Trypsin-EDTA (0.25%) (Thermo Scientific, catalog number: 25200072; store at −20 °C) Pen Strep (Life Technologies, catalog number: 15140–122; store at −20 °C) G-418 Disulfate Salt Solution (Sigma, catalog number: G8168–10ML; store at 4 °C) Forskolin (Sigma, catalog number: F6886–10MG) Glucagon (Sigma, catalog number: G1774-.1MG; store at −20 °C) Demogastrin-2 (DG2) (Pichem, Bat. .. No. M24–061202; store at −20 °C) PYY(3–36) (Sigma, catalog number: P220–500UG; store at −20 °C) Standard Extracellular Saline (SES) Solution, pH 7.4 (see Recipes ) Rat Tail Collagen (RTC) Coating Solution (for one 96-well plate) (see Recipes )
Modification:Article Title: FRET Reporter Assays for cAMP and Calcium in a 96-well Format Using Genetically Encoded Biosensors Expressed in Living Cells
Article Snippet: .. 10 ml Serological Pipet (Krackeler, catalog number: 229010B) 5 ml Serological Pipet (Krackeler, catalog number: 229005B) 1.5 ml Centrifuge Tube (Axygen, catalog number: MCT-150-C) 15 ml Centrifuge Tube (Krackeler, catalog number: 229411) 50 ml Centrifuge Tube (Krackeler, catalog number: 229421) 100 mm x 20 mm Tissue Culture Dish (Krackeler, catalog number: 229621) 96-Well Plate Black, Clear Bottom (Corning, catalog number: 3603) 96-Well Plate, V-Bottom (Greiner Bio-One, catalog number: 651201) FlexStation Pipette Tips (Black, 96) (Molecular Devices, catalog number: 9000–0911) Finntip Flex 300 μl Sterile Pipette Tips (Thermo Scientific, catalog number: 94060513) Channelmate 25 ml Inserts (USA Scientific, catalog number: 1346–2510) Multiwell Plate Washer/dispenser Manifold (Sigma, catalog number: M2781–1EA) 0.2 μm PES Membrane Filter for Vacuum Filtration HEK293/hNPY2R/H188 c18 cells expressing human NPY2R receptor and H188 HEK293/hGCGR/H188 c10 cells expressing human glucagon receptor and H188 HEK293/hCCK 2 R c29 cells expressing human CCK 2 R only Sodium Chloride (Fisher Scientific, catalog number: BP358–1) Sodium Hydroxide (Fisher Scientific, catalog number: BP359–500) Potassium Chloride (Fisher Scientific, catalog number: BP366–500) Magnesium Chloride Hexahydrate (Fisher Scientific, catalog number: BP214–500) Calcium Chloride Dihydrate (Fisher Scientific, catalog number: BP510–100) D-Glucose (Sigma, catalog number: G5767–500G) HEPES 1 M Solution (Sigma, catalog number: H0887; store at 4 °C) Bovine Serum Albumin (Sigma, catalog number: A7030–100G; store at 4 °C) Rat Tail Collagen (Life Technologies, catalog number: A10483–01; store at 4 °C) Glacial Acetic Acid (Thermo Scientific, catalog number: A38S-500) Phosphate-Buffered Saline, 1x (Corning, catalog number: 21–040-CV) DMSO (Sigma, catalog number: D8418–100ML) Fetal Bovine Serum (Sigma, catalog number: 12303C-500ML; store at −20 °C) Dulbecco’s Modified Eagle Medium (Sigma, catalog number: D6429–500ML; store at 4 °C) Trypsin-EDTA (0.25%) (Thermo Scientific, catalog number: 25200072; store at −20 °C) Pen Strep (Life Technologies, catalog number: 15140–122; store at −20 °C) G-418 Disulfate Salt Solution (Sigma, catalog number: G8168–10ML; store at 4 °C) Forskolin (Sigma, catalog number: F6886–10MG) Glucagon (Sigma, catalog number: G1774-.1MG; store at −20 °C) Demogastrin-2 (DG2) (Pichem, Bat. .. No. M24–061202; store at −20 °C) PYY(3–36) (Sigma, catalog number: P220–500UG; store at −20 °C) Standard Extracellular Saline (SES) Solution, pH 7.4 (see Recipes ) Rat Tail Collagen (RTC) Coating Solution (for one 96-well plate) (see Recipes )
Blocking Assay:Article Title: Pyrrolopyrimidine and pyrrolopyridine derivatives
Article Snippet: .. RPMI 1640 (Lonza Cat. No. BE12-115F/U1) Dimethyl sulfoxide (DMSO) (Sigma, Cat. No. D2650) Lipopolysaccharide from E. coli (LPS) (Sigma, Cat. No. L4391) Microplate, 96 well, pp, v-bottom, clear (Greiner; Cat. No. 651201) Master block 96 well, 2 ml (Greiner, Cat. No. 780271) Microplate, 96 well, ps, u-bottom, clear, with lid (Greiner, Cat. No. 650180) 50 mL falcon tube (TPP, Cat. No. 91050) Immulon 2HB 96-well plate (Thermo Fisher Scientific, Cat. No. 3455) Sucrose (Kemika, Cat. No. 1800408) Streptavidin-HRP (Calbiochem, Cat. No. OR03L) Sulphuric acid (Kemika, Cat. No. 1816501) anti-hCCL2 antibody (R&D Systems, Cat. No. MAB679), dissolved in 1 ml of PBS anti-hCCL2 detection antibody (R&D Systems, Cat. No. BAF279), dissolved in 1 ml of PBS Recombinant hCCL2 (standard, R&D Systems, Cat. No. 366-6C) Wash buffer (PBS+0.05% Tween-20) PBS (Sigma, Cat. No. P4417) Tween-20 (Sigma, Cat. No. P2287) Substrate for 20 ml: 18 ml H2O+2 ml 1M sodium acetate+200 μl TMB mix+2.5 μl 30% H2O2 Sodium acetate (Kemika, Cat. No. 1441908) TMB mix (Sigma, Cat. No. T2885) Hydrogen peroxide (Merck, Cat. No. 1.08597) Coating buffer (15 mM Na2CO3 (×H2O)+35 mM NaHCO3) Sodium carbonate (Sigma, Cat. No. S-2127) Sodium bicarbonate (Kemika, Cat. No. 1411007) Assay buffer (PBS+0.05% Tween-20+1% BSA) Bovine Serum Albumin (BSA) (Sigma, Cat. No. A2153) ..
Recombinant:Article Title: Pyrrolopyrimidine and pyrrolopyridine derivatives
Article Snippet: .. RPMI 1640 (Lonza Cat. No. BE12-115F/U1) Dimethyl sulfoxide (DMSO) (Sigma, Cat. No. D2650) Lipopolysaccharide from E. coli (LPS) (Sigma, Cat. No. L4391) Microplate, 96 well, pp, v-bottom, clear (Greiner; Cat. No. 651201) Master block 96 well, 2 ml (Greiner, Cat. No. 780271) Microplate, 96 well, ps, u-bottom, clear, with lid (Greiner, Cat. No. 650180) 50 mL falcon tube (TPP, Cat. No. 91050) Immulon 2HB 96-well plate (Thermo Fisher Scientific, Cat. No. 3455) Sucrose (Kemika, Cat. No. 1800408) Streptavidin-HRP (Calbiochem, Cat. No. OR03L) Sulphuric acid (Kemika, Cat. No. 1816501) anti-hCCL2 antibody (R&D Systems, Cat. No. MAB679), dissolved in 1 ml of PBS anti-hCCL2 detection antibody (R&D Systems, Cat. No. BAF279), dissolved in 1 ml of PBS Recombinant hCCL2 (standard, R&D Systems, Cat. No. 366-6C) Wash buffer (PBS+0.05% Tween-20) PBS (Sigma, Cat. No. P4417) Tween-20 (Sigma, Cat. No. P2287) Substrate for 20 ml: 18 ml H2O+2 ml 1M sodium acetate+200 μl TMB mix+2.5 μl 30% H2O2 Sodium acetate (Kemika, Cat. No. 1441908) TMB mix (Sigma, Cat. No. T2885) Hydrogen peroxide (Merck, Cat. No. 1.08597) Coating buffer (15 mM Na2CO3 (×H2O)+35 mM NaHCO3) Sodium carbonate (Sigma, Cat. No. S-2127) Sodium bicarbonate (Kemika, Cat. No. 1411007) Assay buffer (PBS+0.05% Tween-20+1% BSA) Bovine Serum Albumin (BSA) (Sigma, Cat. No. A2153) ..
other:
High Performance Liquid Chromatography:Article Title: Evaluation of Multiple Methods for Quantification of Glycosaminoglycan Biomarkers in Newborn Dried Blood Spots from Patients with Severe and Attenuated Mucopolysaccharidosis-I
Article Snippet: .. Samples: Donated anonymous healthy newborn dried blood spots (DBS) (from Dr. Francyne Kubaski, Universidade Federal do Rio Grande, Brazil and Dr. Zoltan Lukacs, University of Hamburg, Germany) Donated anonymous MPS-I newborn dried blood spots (DBS) (from the National MPS Society, United States) Standards and Reagents: Internal Standard (IS), “Chondrosine” 2-Amino-2-deoxy-3-O-(b-D-glucopyranuronosyl)-D-galactopyranose (CAS 499-14-9) (Carbosynth, Ref: OA10113) Chondroitin/Dermatan Sulfate Disaccharide “D0a4” (CAS 136144-56-4) (Iduron, Ref: CD002) Heparan Disaccharides Mixture “D0A0” and “D0S0,” I mL of 75 nM (Iduron, Ref: HD Mix) Chondroitinase B from Flavobacterium heparinum, 50 UN (Sigma, Ref: C8058-50UN, 1UN* = 0.1 μmole/hr) Heparinase I from Flavobacterium heparinum, 0.5 IU, lyophilized (Ibex, Ref: 60-400, 1IU* = 1.0 μmole/min) Heparinase II from Flavobacterium heparinum, 0.5 IU, lyophilized (Ibex, Ref: 60-400, 1IU* = 1.0 μmole/min) Heparinase III from Flavobacterium heparinum, 0.5 IU, lyophilized (Ibex, Ref: 60-400, 1IU* = 1.0 μmole/min) Bovine Serium Albumin (BSA) lyophilized powder (Sigma, Ref: A0281-10G) TRIS Ultra-Pure Grade (Amresco, Ref: 201-064-4) Dithiothreitol (DTT) (CAS 3483-12-3) (Sigma Aldrich, Ref: D0632) Calcium chloride (CAS 7440-70-2) (Sigma Aldrich, Ref: C-3306) Methanol, HPLC Grade (CAS 67-56-1) (Fisher, Ref: A452-1) Ammonia Solution 28-30% (CAS 1336-21-6) (MilliporeSigma Ref: 105423) Methanol, Optima Grade (CAS 67-56-1) (Fisher, Ref: A456-4) Water, Optima Grade (CAS 7732-18-5) (Fisher, Ref: W6500) * 1.0 International Unit (IU) = 600 “Sigma” Units (UN) (check definition of units on company spec sheets to be sure) Benchtop Supplies: Deep-well 96-well plate, polypropylene, round bottom, 1mL well (Costar, Ref: 3959) Sealing film for 96-well plate (Axygen, Ref: PCRSP) MilliQ Water Filtration System N2 stream evaporator with 96-well plate adapter (made in-house) Allegra X-12R Centrifuge (Beckman Coulter) Mass Spectrometry Supplies/Equipment: Shallow-well 96-well plate, polypropylene, v-bottom, 350 μL well (Greiner Bio-One, Ref: 651201) Hypercarb porous graphitic carbon column, 50 x 2.1 mm, 5 μm (Thermo, Ref: 35005-052130) Hypercarb guard columns, 10 x 2.1 mm, 5 μm (Thermo, Ref: 35005-012101) Universal uniguard pre-column holder (Thermo, Ref: 852-00) Waters Xevo TQ-S Mass Spectrometer with Water’s I-Class Acquity UPLC with flow-through needle injector 11 ..
Filtration:Article Title: Evaluation of Multiple Methods for Quantification of Glycosaminoglycan Biomarkers in Newborn Dried Blood Spots from Patients with Severe and Attenuated Mucopolysaccharidosis-I
Article Snippet: .. Samples: Donated anonymous healthy newborn dried blood spots (DBS) (from Dr. Francyne Kubaski, Universidade Federal do Rio Grande, Brazil and Dr. Zoltan Lukacs, University of Hamburg, Germany) Donated anonymous MPS-I newborn dried blood spots (DBS) (from the National MPS Society, United States) Standards and Reagents: Internal Standard (IS), “Chondrosine” 2-Amino-2-deoxy-3-O-(b-D-glucopyranuronosyl)-D-galactopyranose (CAS 499-14-9) (Carbosynth, Ref: OA10113) Chondroitin/Dermatan Sulfate Disaccharide “D0a4” (CAS 136144-56-4) (Iduron, Ref: CD002) Heparan Disaccharides Mixture “D0A0” and “D0S0,” I mL of 75 nM (Iduron, Ref: HD Mix) Chondroitinase B from Flavobacterium heparinum, 50 UN (Sigma, Ref: C8058-50UN, 1UN* = 0.1 μmole/hr) Heparinase I from Flavobacterium heparinum, 0.5 IU, lyophilized (Ibex, Ref: 60-400, 1IU* = 1.0 μmole/min) Heparinase II from Flavobacterium heparinum, 0.5 IU, lyophilized (Ibex, Ref: 60-400, 1IU* = 1.0 μmole/min) Heparinase III from Flavobacterium heparinum, 0.5 IU, lyophilized (Ibex, Ref: 60-400, 1IU* = 1.0 μmole/min) Bovine Serium Albumin (BSA) lyophilized powder (Sigma, Ref: A0281-10G) TRIS Ultra-Pure Grade (Amresco, Ref: 201-064-4) Dithiothreitol (DTT) (CAS 3483-12-3) (Sigma Aldrich, Ref: D0632) Calcium chloride (CAS 7440-70-2) (Sigma Aldrich, Ref: C-3306) Methanol, HPLC Grade (CAS 67-56-1) (Fisher, Ref: A452-1) Ammonia Solution 28-30% (CAS 1336-21-6) (MilliporeSigma Ref: 105423) Methanol, Optima Grade (CAS 67-56-1) (Fisher, Ref: A456-4) Water, Optima Grade (CAS 7732-18-5) (Fisher, Ref: W6500) * 1.0 International Unit (IU) = 600 “Sigma” Units (UN) (check definition of units on company spec sheets to be sure) Benchtop Supplies: Deep-well 96-well plate, polypropylene, round bottom, 1mL well (Costar, Ref: 3959) Sealing film for 96-well plate (Axygen, Ref: PCRSP) MilliQ Water Filtration System N2 stream evaporator with 96-well plate adapter (made in-house) Allegra X-12R Centrifuge (Beckman Coulter) Mass Spectrometry Supplies/Equipment: Shallow-well 96-well plate, polypropylene, v-bottom, 350 μL well (Greiner Bio-One, Ref: 651201) Hypercarb porous graphitic carbon column, 50 x 2.1 mm, 5 μm (Thermo, Ref: 35005-052130) Hypercarb guard columns, 10 x 2.1 mm, 5 μm (Thermo, Ref: 35005-012101) Universal uniguard pre-column holder (Thermo, Ref: 852-00) Waters Xevo TQ-S Mass Spectrometer with Water’s I-Class Acquity UPLC with flow-through needle injector 11 ..
Mass Spectrometry:Article Title: Evaluation of Multiple Methods for Quantification of Glycosaminoglycan Biomarkers in Newborn Dried Blood Spots from Patients with Severe and Attenuated Mucopolysaccharidosis-I
Article Snippet: .. Samples: Donated anonymous healthy newborn dried blood spots (DBS) (from Dr. Francyne Kubaski, Universidade Federal do Rio Grande, Brazil and Dr. Zoltan Lukacs, University of Hamburg, Germany) Donated anonymous MPS-I newborn dried blood spots (DBS) (from the National MPS Society, United States) Standards and Reagents: Internal Standard (IS), “Chondrosine” 2-Amino-2-deoxy-3-O-(b-D-glucopyranuronosyl)-D-galactopyranose (CAS 499-14-9) (Carbosynth, Ref: OA10113) Chondroitin/Dermatan Sulfate Disaccharide “D0a4” (CAS 136144-56-4) (Iduron, Ref: CD002) Heparan Disaccharides Mixture “D0A0” and “D0S0,” I mL of 75 nM (Iduron, Ref: HD Mix) Chondroitinase B from Flavobacterium heparinum, 50 UN (Sigma, Ref: C8058-50UN, 1UN* = 0.1 μmole/hr) Heparinase I from Flavobacterium heparinum, 0.5 IU, lyophilized (Ibex, Ref: 60-400, 1IU* = 1.0 μmole/min) Heparinase II from Flavobacterium heparinum, 0.5 IU, lyophilized (Ibex, Ref: 60-400, 1IU* = 1.0 μmole/min) Heparinase III from Flavobacterium heparinum, 0.5 IU, lyophilized (Ibex, Ref: 60-400, 1IU* = 1.0 μmole/min) Bovine Serium Albumin (BSA) lyophilized powder (Sigma, Ref: A0281-10G) TRIS Ultra-Pure Grade (Amresco, Ref: 201-064-4) Dithiothreitol (DTT) (CAS 3483-12-3) (Sigma Aldrich, Ref: D0632) Calcium chloride (CAS 7440-70-2) (Sigma Aldrich, Ref: C-3306) Methanol, HPLC Grade (CAS 67-56-1) (Fisher, Ref: A452-1) Ammonia Solution 28-30% (CAS 1336-21-6) (MilliporeSigma Ref: 105423) Methanol, Optima Grade (CAS 67-56-1) (Fisher, Ref: A456-4) Water, Optima Grade (CAS 7732-18-5) (Fisher, Ref: W6500) * 1.0 International Unit (IU) = 600 “Sigma” Units (UN) (check definition of units on company spec sheets to be sure) Benchtop Supplies: Deep-well 96-well plate, polypropylene, round bottom, 1mL well (Costar, Ref: 3959) Sealing film for 96-well plate (Axygen, Ref: PCRSP) MilliQ Water Filtration System N2 stream evaporator with 96-well plate adapter (made in-house) Allegra X-12R Centrifuge (Beckman Coulter) Mass Spectrometry Supplies/Equipment: Shallow-well 96-well plate, polypropylene, v-bottom, 350 μL well (Greiner Bio-One, Ref: 651201) Hypercarb porous graphitic carbon column, 50 x 2.1 mm, 5 μm (Thermo, Ref: 35005-052130) Hypercarb guard columns, 10 x 2.1 mm, 5 μm (Thermo, Ref: 35005-012101) Universal uniguard pre-column holder (Thermo, Ref: 852-00) Waters Xevo TQ-S Mass Spectrometer with Water’s I-Class Acquity UPLC with flow-through needle injector 11 ..
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