To generate ‘spiked’ SAGM medium for mouse lung alveolar organoids, SABM Small Airway Epithelial Cell Growth Basal Medium (Lonza, CC-3119) was combined with the following additives: SAGM Small Airway Epithelial Cell Growth Medium SingleQuots Supplements and Growth Factors (using only the BPE [2 mL], Insulin [0.5 mL], Retinoic Acid [0.5 mL], Transferrin [0.5 mL], and hEGF [0.5 mL] aliquots) (Lonza, CC-4124), Heat Inactivated Fetal Bovine Serum (Corning, 35-011-CV, final concentration 5%), Antibiotic-Antimycotic (Gibco, 15240-062, final concentration 1x), Cholera Toxin from Vibrio cholerae (Sigma, C8052, final concentration 25 ng/mL).
Cholera toxin from vibrio cholerae
Manufactured by Merck Group
Sourced in Portugal, Germany, Brazil
Cholera Toxin from Vibrio cholerae is a laboratory product derived from the bacterium Vibrio cholerae. It is a protein complex that functions as an exotoxin, disrupting cellular processes within host organisms. This product is primarily used for research purposes in various scientific fields.
Automatically generated - may contain errors
Lab products found in correlation
Fetal bovine serum (fbs), by Thermo Fisher Scientific (3 mentions)
Hydrocortisone, by Merck Group (2 mentions)
Fetal bovine serum (fbs), by Merck Group (2 mentions)
Antibiotic antimycotic, by Thermo Fisher Scientific (1 mentions)
Fetal bovine serum (fbs), by Corning (1 mentions)
Dimethyl sulfoxide (dmso), by Thermo Fisher Scientific (1 mentions)
Dulbecco phosphate buffered saline (dpbs), by Thermo Fisher Scientific (1 mentions)
Mccoy s 5a, by Thermo Fisher Scientific (1 mentions)
Rpmi 1640, by Thermo Fisher Scientific (1 mentions)
Trypsin edta, by Thermo Fisher Scientific (1 mentions)
Dmem f12, by Thermo Fisher Scientific (1 mentions)
Medium 199, by Thermo Fisher Scientific (1 mentions)
Mcdb 105, by Cell Applications (1 mentions)
Bovine insulin, by Merck Group (1 mentions)
Arium pro water purification system, by Sartorius (1 mentions)
Insulin, by Thermo Fisher Scientific (1 mentions)
Fetal bovine serum (fbs), by GE Healthcare (1 mentions)
F 12k medium, by Thermo Fisher Scientific (1 mentions)
Insulin, by Merck Group (1 mentions)
Streptomycin, by Merck Group (1 mentions)
6 protocols using cholera toxin from vibrio cholerae
1
Mouse Lung Alveolar Organoid Culture
2
Optimized Cell Culture Conditions
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McCoy’s 5A (#16600-082), Medium 199 (#11150-059), RPMI1640 (#11875-085), DMEM/F12 (#11320-033), 1% Penn Strep (Penicillin/Streptomycin; P/S, #15140122), MEM Non-Essential Amino Acids (NEAA) Solution (100X, #11140-050), DPBS (#14040-117), and Trypsin-EDTA (0.25%, #25200-056) were purchased from Gibco (Life Technologies, Grand Island, NY). MCDB105 (#117-500) was purchased from Cell Applications Inc. (San Diego, CA). Alpha-MEM was obtained from Irvine Scientific (Irvine, CA) OCMI-E was purchased from University of Miami (Live Tumor Culture Core at Sylvester Comprehensive Cancer Center, University of Miami Miller School of Medicine, Miami, FL). Fetal bovine serum (FBS, #F2442) and Cholera Toxin from Vibrio cholerae (#C8052) were purchased from Sigma-Aldrich (St. Louis, MO). Dimethyl Sulfoxide (DMSO, #CAS 67-68-5) was purchased from Thermo Fisher Scientific (Waltham, MA).
3
Establishment of 3D Organotypic Cultures
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Cisplatin (cis-dichlorodiammine platinum(II), 99.9%), potassium tetrachloropalladate (II) (K2PdCl4, 98%), spermine (N,N’-bis(3-aminopropyl)-1,4-diaminobutane, 99%), Dulbecco’s Modified Eagle Medium—high-glucose cell growth medium (DMEM-HG), 1:1 mixture of Dulbecco’s Modified Eagle Medium and Ham’s F12 cell growth medium (DMEM/F12 1:1), human epidermal growth factor (hEGF; recombinant, expressed in E. coli), cholera toxin from Vibrio cholerae, bovine insulin (10 mg/mL insulin in 25 mM HEPES, pH 8.2) and hydrocortisone were purchased from Sigma-Aldrich (Sintra, Portugal). Fetal bovine serum (FBS) and horse serum were from Gibco (Thermo Fisher Scientific, Inc., Waltham, MA, USA). Ultrapure water (18.2 MΩ × cm at 25 °C) was obtained with an arium® pro water purification system (Sartorius, Goettingen, Germany). Animals were anesthetized with isoflurane inhalation (IsoFlo®, 100% isoflurane) acquired from Abbott (Berkshire, UK). All the other reagents were of analytical grade.
4
Intoxication of CaCo-2 Cells by CDTa and CDTb
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The plasmid pEGFP-N1/LSR was used for the ectopic expression of LSR-EGFP in CaCo-2 cells and is described elsewhere [27 (link)].
CDTa and the precursor form of CDTb (pCDTb) were purified as recombinant proteins from the expression host Bacillus megaterium as described earlier [28 (link)]. CDTb was obtained by proteolytic activation of pCDTb [29 (link), 30 (link)]. Intoxication experiments with CaCo-2 cells were performed by adding 4 nM CDTa together with 5 nM CDTb directly to the cell culture medium.
Recombinant production and DyLight488-labeling of the receptor-binding domain (RBD) are described elsewhere [27 (link)]. Cholera toxin from Vibrio cholerae was obtained from Sigma-Aldrich (C8052).
CDTa and the precursor form of CDTb (pCDTb) were purified as recombinant proteins from the expression host Bacillus megaterium as described earlier [28 (link)]. CDTb was obtained by proteolytic activation of pCDTb [29 (link), 30 (link)]. Intoxication experiments with CaCo-2 cells were performed by adding 4 nM CDTa together with 5 nM CDTb directly to the cell culture medium.
Recombinant production and DyLight488-labeling of the receptor-binding domain (RBD) are described elsewhere [27 (link)]. Cholera toxin from Vibrio cholerae was obtained from Sigma-Aldrich (C8052).
5
Cell Culture Conditions for Cancer Research
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MV4-11 cells were cultured in the RPMI 1640 medium (HIIET PAS, Poland) supplemented with 1.0 mM sodium pyruvate, 2 mM L-glutamine and 10% FBS (all from Sigma Aldrich, Steinheim, Germany), LoVo cells were cultured in the F-12K medium (Life Technologies, Carlsbad, CA, United States) supplemented with 10% FBS (GE Healthcare, Chicago, IL, USA), and LoVo/DX cells were cultured in the mixture of RPMI 1640+OptiMEM (1:1) medium (HIIET PAS) supplemented with 5% FBS (GE Healthcare, Chicago, IL, USA), 2 mM L-glutamine, 1.0 mM sodium pyruvate (all from Sigma Aldrich, Steinheim, Germany) and 0.1 µg/mL doxorubicin chloride (Accord).
MCF-7 cells were cultured in the Eagle’s medium (HIIET PAS, Poland) supplemented with 10% FBS, 8 µg/mL insulin, 2 mM L-glutamine and 1% MEM-non essential amino acid solution 100X (all from Sigma Aldrich, Steinheim, Germany). MCF-10A cells were cultured in the HAM’S F-12 with L-glutamine medium (Corning) supplemented with 10% Hors Serum (Gibco), 10 µg/mL insulin, 0.5 µg/mL hydrocortisone, 20 ng/mL EGFh and 0.05 mg/mL Cholera Toxin from Vibrio cholerae (all from Sigma Aldrich, Steinheim, Germany). All culture mediums were supplemented with 100 units/mL penicillin (Polfa Tarchomin S.A. Poland), and 100 µg/mL streptomycin (Sigma Aldrich). All cell lines were grown at 37 °C with 5% CO2 humidified atmosphere.
MCF-7 cells were cultured in the Eagle’s medium (HIIET PAS, Poland) supplemented with 10% FBS, 8 µg/mL insulin, 2 mM L-glutamine and 1% MEM-non essential amino acid solution 100X (all from Sigma Aldrich, Steinheim, Germany). MCF-10A cells were cultured in the HAM’S F-12 with L-glutamine medium (Corning) supplemented with 10% Hors Serum (Gibco), 10 µg/mL insulin, 0.5 µg/mL hydrocortisone, 20 ng/mL EGFh and 0.05 mg/mL Cholera Toxin from Vibrio cholerae (all from Sigma Aldrich, Steinheim, Germany). All culture mediums were supplemented with 100 units/mL penicillin (Polfa Tarchomin S.A. Poland), and 100 µg/mL streptomycin (Sigma Aldrich). All cell lines were grown at 37 °C with 5% CO2 humidified atmosphere.
6
Proteomic Analysis of Bacterial Toxins
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Acrylamide (17-1302-02), bis-Acrylamide (17-1304-02), sodium dodecyl sulphate (SDS) (17-1313-01), HiTrap Protein-A Sepharose high-performance column (17-0402-01), and Superdex 75 Increase 10/300 GL (29148721) were acquired from GE Healthcare Life Sciences (São Paulo, SP, Brazil) . Azocasein (A2765), bromelain (code B4882), Lcysteine (C7352), O-phthaldialdehyde (P0657), β-mercaptoethanol (M6250), Evans blue dye (E2129), cholera toxin from Vibrio cholerae (C8052), Freund's complete (F5881) and incomplete (F5506) adjuvant, alkaline phosphatase-conjugated goat anti-rabbit IgG antibodies (A6066) and p-nitrophenylphosphate disodium (N2765) were obtained from Sigma-Aldrich (São Paulo, SP, Brazil). All other chemicals were of analytical grade.
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