General reagents were purchased from Sigma-Aldrich (St. Louis, MO). High-performance liquid chromatography (HPLC)-grade water and acetonitrile, Gibco phosphate buffered saline (PBS), and Gibco Iscove's Modified Dulbecco's Medium (IMDM) were purchased from Thermo Fisher Scientific (Waltham, MA); ammonium formate (Alfa Aesar, 99% purity) from VWR International (Radnor, PA); S-adenosyl-L-methionine (AdoMet) sulfate p-toluenesulfonate salt from Affymetrix (Santa Clara, CA); S-adenosyl-L-[carboxy-14C]-methionine (14C-AdoMet) from American Radiolabeled Chemicals (St. Louis, MO); Fetal Bovine Serum (FBS) from Atlanta Biologicals (Flowery Branch, GA); Chicken Serum from Gemini Bio-Products (West Sacramento, CA).
Gibco phosphate buffered saline
Manufactured by Thermo Fisher Scientific
Sourced in United States
Gibco Phosphate Buffered Saline (PBS) is a commonly used buffer solution that maintains a physiological pH. It is a sterile, isotonic solution that helps maintain the ionic balance and osmotic pressure of cells and tissues. PBS is widely used in various laboratory applications, such as cell culture, immunoassays, and sample preparation.
Automatically generated - may contain errors
Lab products found in correlation
3 4 5 dimethylthiazol 2 yl 2 5 diphenyltetrazolium bromide mtt, by Merck Group (2 mentions)
Dimethyl sulfoxide (dmso), by Merck Group (2 mentions)
Fetal bovine serum (fbs), by Atlanta Biologicals (1 mentions)
Ammonium formate, by Avantor (1 mentions)
Chicken serum, by Gemini Bio (1 mentions)
General reagents, by Merck Group (1 mentions)
S adenosyl l methionine adomet sulfate p toluenesulfonate salt, by Thermo Fisher Scientific (1 mentions)
S adenosyl l carboxy 14c methionine 14c adomet, by American Radiolabeled Chemicals (1 mentions)
Odyssey imaging system, by LI COR (1 mentions)
Enhanced chemiluminescence kit, by Cytiva (1 mentions)
Mouse anti human gapdh monoclonal antibody, by Cell Signaling Technology (1 mentions)
Dabrafenib, by Merck Group (1 mentions)
Ab2074, by Abcam (1 mentions)
Phospho akt ser473, by Cell Signaling Technology (1 mentions)
Dabrafenib, by Selleck Chemicals (1 mentions)
Dabrafenib, by Active Motif (1 mentions)
Sodium dodecyl sulphate polyacrylamide gel electrophoresis, by Bio-Rad (1 mentions)
Sc 9104, by Santa Cruz Biotechnology (1 mentions)
Superdex 200 increase 10 300 gl column, by GE Healthcare (1 mentions)
Mabselect sure lx resin, by GE Healthcare (1 mentions)
8 protocols using gibco phosphate buffered saline
1
Reagents for Cell Culture Studies
2
Western Blot Analysis of IDO Protein
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
The treated cells were washed twice with cold Gibco® phosphate-buffered saline (Thermo Fisher Scientific) and were lysed in RIPA buffer (Biocolor BioScience & Technology Co., Ltd., Shanghai, China) in the presence of a proteinase inhibitor (Kangchen Bio-tech, Shanghai, China). Protein quantification was performed with the BCA-100 protein assay kit (Biocolor BioScience & Technology Co., Ltd.). Samples were subjected to 8% sodium dodecyl sulphate polyacrylamide gel electrophoresis, transferred to polyvinylidene fluoride (PVDF) membranes (EMD Millipore, Billerica, MA, USA) and blocked with 5% fat-free milk for 2 h at room temperature. Equal loading in each blot was confirmed by Coomassie staining (Beyotime Biotechnology, Shanghai, China) of the membrane. The membrane was incubated overnight with a mouse anti-human IDO monoclonal antibody (1:500; Abcam, Cambridge, MA, USA) and mouse anti-human GAPDH monoclonal antibody (1:5,000; Cell Signaling Technology, Inc., Beverly, MA, USA) at 4°C. Then, the membrane was incubated with the horseradish peroxidase (HRP)-labeled secondary antibody (1:7,000) for 1 h at room temperature. Signals were visualized with an enhanced chemiluminescence kit (Amersham Pharmacia Biotech, Piscataway, NJ, USA) and quantified using an Odyssey® imaging system (LI-COR Inc., Lincoln, NE, USA). The protein level of IDO was normalized to the level of GAPDH.
3
Dabrafenib Treatment and Protein Expression
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Dabrafenib was purchased from Selleckchem (Houston, TX) and dissolved in dimethyl sulfoxide (DMSO) (Sigma-Aldrich) at a final concentration of 1.92 mM. Dabrafenib was stored as stock solutions at -80°C and diluted in CM just before use. 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) was purchased from Sigma-Aldrich, dissolved at a concentration of 5 mg/ml in GIBCO™ Phosphate-Buffered Saline (PBS) (Invitrogen, Thermo Fisher Scientific, Waltham, MA) and stored at 4°C. Mouse monoclonal antibodies (mAb) against ADAM9 (sc-377233), and rabbit polyclonal antibody against human β-tubulin (sc-9104) were purchased from Santa Cruz Biotechnology, Inc. (Santa Cruz, CA). mAb against PIK3R2 (#05-217) was purchased from Upstate Biotechnology (Lake Placid, NY). mAb against MMP7 (#111433) was purchased from R&D Systems (Minneapolis, MN). Rabbit polyclonal antibody against CXCR4 (ab2074) was purchased from Abcam (Cambridge, MA). Rabbit polyclonal antibodies against ERK1/2 (#9102), phospho-ERK1/2 (Thr202/Tyr204) (#9101), AKT (#9272) and phospho-AKT (Ser473) (#9271) were purchased from Cell Signaling Technology, Inc. (Beverly, MA). Reagents for sodium dodecyl sulphate (SDS)-polyacrylamide gel electrophoresis were all purchased from Bio-Rad Laboratories, Inc. (Hercules, CA).
4
Cytotoxicity Assay for Cancer Cell Lines
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Dabrafenib (GSK2118436A) and LEE011 (Active Biochem, Hong Kong) were dissolved in DMSO (Sigma-Aldrich) at a final concentration of 1.92 mM and 16 mM, respectively. Drugs were stored as stock solutions at –80°C and diluted in CM just before use.
3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) was purchased from Sigma-Aldrich, dissolved at a concentration of 5 mg/ml in GIBCO™ Phosphate-Buffered Saline (PBS) (Invitrogen, Thermo Fisher Scientific, Waltham, MA) and stored at 4°C.
Mouse monoclonal antibody (mAb) against PTTG1 (DCS-280) and rabbit polyclonal antibody against human β-tubulin (sc-9104) were purchased from Santa Cruz Biotechnology, Inc., (Santa Cruz, CA).
Reagents for sodium dodecyl sulphate (SDS)-polyacrylamide gel electrophoresis were all purchased from Bio-Rad Laboratories, Inc. (Hercules, CA).
3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) was purchased from Sigma-Aldrich, dissolved at a concentration of 5 mg/ml in GIBCO™ Phosphate-Buffered Saline (PBS) (Invitrogen, Thermo Fisher Scientific, Waltham, MA) and stored at 4°C.
Mouse monoclonal antibody (mAb) against PTTG1 (DCS-280) and rabbit polyclonal antibody against human β-tubulin (sc-9104) were purchased from Santa Cruz Biotechnology, Inc., (Santa Cruz, CA).
Reagents for sodium dodecyl sulphate (SDS)-polyacrylamide gel electrophoresis were all purchased from Bio-Rad Laboratories, Inc. (Hercules, CA).
5
Protein A Affinity Purification of Antibodies
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Cell culture supernatant was incubated with Protein A (ProA) affinity MabSelect SuRe LX resin (GE Healthcare, Pittsburgh, PA, USA) overnight (1 mL resin for 40 mg target antibody or HC dimer estimated by ForteBio Octet) for batch binding. Affinity resin with bound protein was collected by filtration and transferred to a disposable column. Bound resin was washed with 20X column volumes of 1X Gibco Phosphate buffered saline (PBS) (Thermo Fisher Scientific, Waltham, MA, USA) in batch mode. Desired protein was eluted by approximately 5 column volumes of 20 mM sodium acetate pH 3.5 buffer. Eluate was immediately buffer exchanged into 20 mM sodium acetate pH 5.5 buffer using 10K MWCO Slide-A-Lyzer dialysis cassettes. To further polish material for analytical and biophysical characterization efforts (DSC, SEC-MALS, LC-MS), desired product was purified to >98% purity (by SE-UPLC and cSDS) on a Superdex 200 Increase 10/300GL column (GE Healthcare Bio-Science AB, Uppsala, Sweden). Mobile phase was 20 mM sodium acetate, 200 mM sodium chloride pH 5.5 buffer. All purified protein was buffer exchanged overnight using 10K MWCO Slide-A-Lyzer dialysis cassettes into 1X Gibco PBS pH 7.4 and normalized to 1 mg/mL for characterization. Concentration was determined by UV absorbance at 280 nm on a Nanodrop 2000 1- position Spectrophotometer (Thermo Scientific).
6
Nanoparticle Size Analysis via NanoSight NS300
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
All experiments were carried out using a NanoSight NS300 (Malvern PANalytical, Malvern, United Kingdom), with NTA software version 3.2 (Malvern Instruments). All samples were diluted using Gibco phosphate buffered saline (PBS, catalogue number 14190094) (Thermo Fisher, Paisley, United Kingdom) except where otherwise noted. The PBS was filtered prior to use using a Millex 33 mm, 0.22 μm syringe filter unit (Merck, Nottingham, United Kingdom). Samples were vortexed briefly, filtered using a 4 mm, 0.22 μm Millex syringe filter unit (Merck), except where otherwise noted, then taken up into a 1 mL BD Plastipak syringe (BD Biosciences, Wokingham, United Kingdom) and injected into the flow cell of the NanoSight NS300. Once loaded, the sample was passed through the system using a syringe pump at a rate of 50 arbitrary units (A.U.), decreased gradually through 500, 300, 200 and 100 A.U. Once stable, this flow rate allows the particles to pass across the field of view in approximately 8 s. A video would then be captured and processed (script attached, Supplementary File 1 ).
7
Quantifying Anaerobic Fecal Bacteria
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
We suspended fresh fecal pellets in sterile, prereduced Gibco phosphate-buffered saline (PBS; Thermo Fisher) using a ratio of 1 part feces to 9 parts Gibco PBS (wt/vol; Thermo Fisher, Waltham, MA). We serially diluted these suspensions, plated them on TCCFA, and then incubated the plates anaerobically at 37°C for 18 to 24 h before counting the colonies.
8
Immunofluorescent Staining of FFPE Tissues
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Formalin-fixed, paraffin-embedded tissue sections (4 µm) were blocked with a blocking buffer containing 1% normal swine serum blocking solution (Vector Laboratories, Burlingame, CA, USA), 1% bovine serum albumin (BSA) (Sigma-Aldrich, Burlington, MA, USA), and 0.1% gelatin from cold-water fish skin (Sigma-Aldrich) in 1× Gibco phosphate-buffered saline (PBS) (Thermo Fisher Scientific). Sections were incubated overnight with the primary antibodies anti-CK19 (mouse, 1:200, MU246-UC, BioGenex, Fremont, CA, USA) and anti-ALG9 (rabbit, 1:200, HPA038575, Sigma-Aldrich) at 4 °C. Secondary antibodies FITC 490 anti-rabbit (goat, 1:200, 111-005-003, Jackson ImmunoResearch, West Grove, PA, USA) and TexasRed 570 anti-mouse (goat, 1:200, 115-075-062, Jackson ImmunoResearch) were incubated for 45 min at room temperature. Protein expression and localization were visualized with the Zeiss Imager Z2 microscope combined with the ApoTome.2 (Zeiss, Oberkochen, Germany) and analyzed with ZEN 2 blue (version 10, Zeiss).
About PubCompare
Our mission is to provide scientists with the largest repository of trustworthy protocols and intelligent analytical tools, thereby offering them extensive information to design robust protocols aimed at minimizing the risk of failures.
We believe that the most crucial aspect is to grant scientists access to a wide range of reliable sources and new useful tools that surpass human capabilities.
However, we trust in allowing scientists to determine how to construct their own protocols based on this information, as they are the experts in their field.
Ready to get started?
Sign up for free.
Registration takes 20 seconds.
Available from any computer
No download required
Revolutionizing how scientists
search and build protocols!