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Pierce 660 reagent

Manufactured by Thermo Fisher Scientific
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The Pierce 660 reagent is a protein quantification solution designed for determining the concentration of proteins in samples. It operates on the principle of colorimetric detection, providing a rapid and reliable method for measuring protein levels. The reagent can be used with a variety of protein samples and is compatible with common laboratory instruments for absorbance measurement.

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Lab products found in correlation

Imagequant las4000 reader, by GE Healthcare (1 mentions) Tp401, by OriGene (1 mentions) Anti gfp, by OriGene (1 mentions) Anti flag m2, by Merck Group (1 mentions) Hrp conjugated secondary antibody, by Jackson ImmunoResearch (1 mentions) Protease inhibitor cocktail, by Roche (1 mentions) Rabbit anti histone h3, by Cell Signaling Technology (1 mentions)

Close spelling variants of this product

Pierce 660 nm protein assay reagent kit Pierce 660 nm reagent Pierce 660 nm Protein Assay Reagent Pierce 660 nm Protein Assay with Ionic detergent compatibility reagent Pierce-660 nm ready-to-use Protein Assay Reagent Pierce 660 nm Assay Reagent Pierce 660 Protein Assay Reagent Pierce reagents

3 protocols using pierce 660 reagent

1

Immunoblotting of Flag, GFP, and α-Spectrin

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S2 cells were lysed in 50 mM Tris-HCl (pH 7.8), 150 mM NaCl, 1 mM EDTA (pH 8.0), 1% Triton X-100, 0.01% Igepal, and protease inhibitors (Roche Applied Science). Protein concentration was quantified using Pierce 660 reagent (22660, Thermo Scientific) according to manufacturer’s instructions. Following SDS-PAGE, proteins were detected by immunoblotting with anti-Flag M2 (1:1000, Sigma Aldrich), anti-GFP (1:5000, TP401, Acris) and anti-α-Spectrin (1:1000, RRID: AB_528473) antibodies, followed by incubation with corresponding HRP-conjugated secondary antibodies (Jackson Immuno Research). Chemiluminescent signal was captured using ImageQuant LAS4000 reader (GE Healthcare).
Donohoe C.D., Csordás G., Correia A., Jindra M., Klein C., Habermann B, & Uhlirova M. (2018). Atf3 links loss of epithelial polarity to defects in cell differentiation and cytoarchitecture. PLoS Genetics, 14(3), e1007241.
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2

Protein Extraction and Western Blot Analysis

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The antibody details (company, catalog number, species, and dilution) are provided in Table 1. Cells were treated with the drugs at the indicated concentrations, washed with PBS, followed by lysis in 1X RIPA buffer (150mM NaCl, 1% NP-40, 0.5% sodium deoxycholate, 0.1% SDS and 100mM Tris, pH 8.0) containing protease inhibitor cocktail (Roche, Indianapolis, IN), 1mM sodium fluoride, 1mM sodium orthovanadate and 1mM PMSF. In the case of PDAC tissue samples homogenization was carried out using an electric homogenizer in RIPA lysis buffer. Protein estimation was performed against a BSA standard using the Pierce 660 reagent (Thermo Fisher Scientific, Waltham, MA). 20μg protein was loaded on a 12% SDS-PAGE gel and western blot analysis was carried out as described previously [19 ].
Woods N., Trevino J., Coppola D., Chellappan S., Yang S, & Padmanabhan J. (2015). Fendiline inhibits proliferation and invasion of pancreatic cancer cells by interfering with ADAM10 activation and β-catenin signaling. Oncotarget, 6(34), 35931-35948.
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3

Western Blot Analysis Protocol

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Western blots were performed as described by AbCam (http://www.abcam.com/ps/pdf/protocols/WB-beginner.pdf). In brief, cells were lysed in ice-cold RIPA buffer (20 mM Tris-HCl, pH 7.5, 150 mM NaCl, 1 mM EDTA, 1 mM EGTA, 1% NP-40, 1% sodium deoxycholate) supplemented with 1 mM phenylmethylsulfonyl fluoride and 1 mM sodium fluoride, and total protein amount was measured using a Pierce 660 reagent (Thermo Scientific) with bovine serum albumin as a standard. Proteins were separated by SDS-PAGE, and transferred to nitrocellulose membranes. The membrane was blocked with phosphate-buffered saline (PBS) supplemented with 5% bovine serum albumin, and probed with the appropriate primary antibody (rabbit anti-phospho-p42/p44, rabbit anti-p42/p44, rabbit anti-pT11-Histone H3, and rabbit anti-Histone H3 from Cell Signaling; rabbit anti-GAPDH and mouse-anti-Myc from Sigma-Aldrich) overnight at 4°C. After primary antibody incubation, membranes were probed with horseradish peroxidase (HRP) conjugated goat anti-rabbit IgG or goat anti-mouse IgG secondary antibodies (Bio-Rad). HRP-conjugated secondary antibodies were then detected using commercial chemiluminescence substrates (Pierce). Chemiluminescence images were acquired using a FluorChem M FM0455 imager.
Keller K.E., Doctor Z.M., Dwyer Z.W, & Lee Y.S. (2014). SAICAR induces protein kinase activity of PKM2 that is necessary for sustained proliferative signaling of cancer cells. Molecular cell, 53(5), 700-709.
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