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Complete lysis buffer

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Complete lysis buffer is a reagent used in molecular biology applications to disrupt and solubilize cellular components, including proteins, DNA, and RNA. It functions by breaking down the cell membrane and organelles, allowing for the extraction and purification of cellular contents for further analysis or experimentation.

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Lab products found in correlation

Dc protein assay, by Bio-Rad (7 mentions) Protease and phosphatase inhibitor cocktail, by Roche (7 mentions) Gapdh, by Cell Signaling Technology (3 mentions) Anti β actin, by Merck Group (3 mentions) β actin, by Merck Group (2 mentions) N cadherin, by Cell Signaling Technology (2 mentions) Anti β actin a5441, by Merck Group (1 mentions) N cadherin 4061, by Cell Signaling Technology (1 mentions) Claudin 5, by Cell Signaling Technology (1 mentions) Pan akt, by Cell Signaling Technology (1 mentions) Ser33 37 thr41β catenin, by Cell Signaling Technology (1 mentions) Zo 1 and zo 2, by Cell Signaling Technology (1 mentions) T p38 mapk, by Cell Signaling Technology (1 mentions) P src tyr416, by Cell Signaling Technology (1 mentions) T src, by Cell Signaling Technology (1 mentions) P p38 mapk, by Cell Signaling Technology (1 mentions) Claudin 5 antibody, by Abcam (1 mentions) α sma, by Merck Group (1 mentions) Gapdh 2118l, by Cell Signaling Technology (1 mentions) Enos 610297, by BD (1 mentions)

7 protocols using complete lysis buffer

1

Western Blot Analysis of Cell Signaling

Cited 2 times
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Cell lysates were prepared using complete lysis buffer (EMD Millipore, San Diego, CA), with protease and phosphatase inhibitor cocktails (Roche Diagnostics, Indianapolis, IN). Protein quantification was performed using DC protein assay from Bio-Rad (Hercules, CA). Western blot analysis was performed as described previously (Al-Azayzih et al. 2015 (link); Sabbineni et al. 2019 (link)). Antibodies used include Akt1 (2938), N-cadherin (4061), phosphorylated p38-MAPK (9211), total p38-MAPK (9212), phosphorylated Smad2/3 (8828), total Smad2/3 (8685), FoxC2 (12974), phosphorylated β-catenin (9561), total β-catenin (8480) and GAPDH (2118) from Cell Signaling Technology (Danvers, MA). Anti-β-actin (A5441) was from Sigma (St. Louis, MO), anti-eNOS (610297) was from BD Pharmingen (San Diego, CA), anti-ALK-5 (ab31013) was purchased from Abcam (Cambridge, UK) and anti-ALK-1 (NBP1-90254) was obtained from Novus Biologicals. Anti-phosphorylated-Smad1/5/8 (AB3848-I) from Millipore Sigma and anti-Smad1/Smad5/Smad8 (SAB2702532) from Sigma-Aldrich. Band densitometry was performed using NIH Image J software (Bethesda, MD).
Verma A., Artham S, & Somanath P.R. (2020). ALK-1 to ALK-5 ratio dictated by the Akt1-β-catenin pathway regulates TGFβ-induced endothelial-to-mesenchymal transition. Gene, 768, 145293.
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2

Western Blot Analysis of Cell Signaling

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Cell lysates were prepared using complete lysis buffer (EMD Millipore, San Diego, CA) with protease and phosphatase inhibitor cocktails (Roche Diagnostics, Indianapolis, IN). Protein quantification was performed using Dc protein assay (Bio-Rad, Hercules, CA). Western blot analysis was performed as described previously.16 (link), 17 (link) Antibodies used include Akt1, Akt2, Akt3, pan-Akt, Tyr216GSK-3β, Ser33/37/Thr41β-catenin, claudin-5, ZO-1 and ZO-2 (Cell Signaling) and anti-β-actin (Sigma). Densitometry was done using NIH Image J software.
Gao F., Alwhaibi A., Artham S., Verma A, & Somanath P.R. (2018). Endothelial Akt1 loss promotes prostate cancer metastasis via β-catenin-regulated tight-junction protein turnover. British Journal of Cancer, 118(11), 1464-1475.
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3

Western Blotting Protein Analysis

Cited 1 time
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Western analysis was performed as described previously (20 (link)). Cell lysates were prepared using complete lysis buffer (EMD Millipore, San Diego, CA) with protease and phosphatase inhibitor cocktails (Roche Diagnostics, Indianapolis, IN). Protein quantification was performed using DC protein assay from Bio-Rad (Hercules, and CA). Western blot analysis was performed as described previously (21 (link),22 (link)). Antibodies used include stromelysin1 (cat. No. 14351-S) dilution 1:1,000 in milk, vascular endothelial cadherin (VE-cadherins; cat No. 2158) dilution 1:1,000 in BSA, P-P38 MAPK (cat No. 9112-S) dilution 1:1,000 in BSA, T-P38MAPK (cat No. 9212-S) dilution 1:1,000 in BSA, P-SRC Tyr-416 (cat No. 6943-S) dilution 1:1,000 in BSA and T-SRC (cat No. 2109-S) dilution 1:1,000 in BSA all from Cell Signaling Technology (Danvers, MA). β-actin (dilution in milk, primary antibodies 1:10,000 and secondary antibodies 1:20,000) from Sigma (St. Louis, MO) and Claudin-5 antibodies (cat No. ab15106) 1:1,000 and secondary antibodies 1:5,000 dilution in milk from Abcam (Cambridge, MA). Band densitometry was done using NIH Image J software.
Kadry R.W., Adil M.S., Newsome A.S, & Somanath P.R. (2021). Cisatracurium attenuates LPS-induced modulation of MMP3 and junctional protein expression in human microvascular endothelial cells. Bioscience trends, 15(1), 50-54.
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4

Western Blot Analysis of Cell Lysates

Cited 1 time
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Cell lysates were prepared using complete lysis buffer (EMD Millipore, San Diego, CA) with protease and phosphatase inhibitor cocktails (Roche Diagnostics, Indianapolis, IN). Protein quantification was performed using DC protein assay from Bio-Rad (Hercules, and CA). Western blot analysis was performed as described previously (Goc et al., 2013 (link); Sabbineni et al., 2018 (link)). Antibodies used include stromelysin1, N-cadherin, and GAPDH from Cell Signaling Technology (Danvers, MA), αSMA, and β-actin from Sigma (St. Louis, MO), and ALK5 from Abcam (Cambridge, MA). Band densitometry was done using NIH Image J software.
Alharthi A., Verma A., Sabbineni H., Adil M.S, & Somanath P.R. (2020). Distinct effects of pharmacological inhibition of stromelysin1 on endothelial-to-mesenchymal transition and myofibroblast differentiation. Journal of cellular physiology, 236(7), 5147-5161.
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5

Western Blot Analysis of Epithelial-Mesenchymal Markers

Cited 1 time
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Cell lysates were prepared using complete lysis buffer (EMD Millipore, San Diego, CA) with protease and phosphatase inhibitor cocktails (Roche Diagnostics, Indianapolis, IN). Protein quantification was performed using DC protein assay from Bio-Rad (Hercules, and CA). Western blot analysis was performed as described previously (Abdalla et al., 2013 (link); Al-Azayzih et al., 2015 (link)). Antibodies used include N-cadherin (4061), VE-cadherin (2158S), phosphorylated p-38 MAPK (9211S), total p38-MAPK (9212S), phosphorylated Smad2/3 (8828S), total Smad2/3 (8685S), FoxC2 (12974S), Snail (3879S), and GAPDH (2118L) from Cell Signaling Technology (Danvers, MA), αSMA (A2547) and β-actin (A5441) from Sigma (St. Louis, MO), eNOS (610297) from BD Pharmingen (San Diego, CA), and TGFβ2 (MAB612) from R&D (Minneapolis, MN). Band densitometry was done using NIH Image J software.
Sabbineni H., Verma A, & Somanath P.R. (2018). Isoform-Specific Effects of Transforming Growth Factor-β on Endothelial to Mesenchymal Transition. Journal of cellular physiology, 233(11), 8418-8428.
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6

Protein Expression Analysis by Western Blot

Cited 1 time
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Cell lysates were prepared using complete lysis buffer (EMD Millipore, San Diego, CA) with protease and phosphatase inhibitor cocktails (Roche Diagnostics, Indianapolis, IN). Protein quantification was performed using the DC protein assay (Bio-Rad, Hercules, CA). Western blot analysis was performed as described previously [37 (link), 38 (link)]. Antibodies used include Akt1, β-catenin (ser675), N-cadherin, VE-cadherin, Snail1, FoxC2, phospho- and total-Smad 2/3, phospho- and total-p-38 MAPK, GAPDH from Cell Signaling (Danvers, MA)., anti- β-actin, alpha-SMA, and TGFβ2 from Sigma Aldrich (St. Louis, MO) and eNOS from BD Pharmingen (SanDiego, CA). HRP-conjugated goat-anti-mouse and goat-anti-rabbit secondary antibodies were obtained from Bio-Rad (Hercules, CA). Densitometry was done using NIH Image J software.
Sabbineni H., Verma A., Artham S., Anderson D., Amaka O., Liu F., Narayanan S.P, & Somanath P.R. (2019). Pharmacological inhibition of β-catenin prevents EndMT in vitro and vascular remodeling in vivo resulting from endothelial Akt1 suppression. Biochemical pharmacology, 164, 205-215.
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7

Western Blot Analysis of Signaling Proteins

Cited 1 time
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Cell lysates were prepared using complete lysis buffer (EMD Millipore, San Diego, CA) with protease and phosphatase inhibitor cocktails (Roche Diagnostics, Indianapolis, IN). Protein quantification was performed using DC protein assay (Bio-Rad, Hercules, CA. Western blot analysis was performed as described previously (Goc, Al-Azayzih et al. 2013 (link), Gao, Al-Azayzih et al. 2015 (link)). Antibodies used include Src Tyr416, total-Src, Akt Ser473, total-Akt (Cell Signaling, Danvers, MA) and anti- β-actin (Sigma, St. Louis, MO). Densitometry was done using NIH Image J software.
Gao F., Sabbineni H., Artham S, & Somanath P.R. (2017). Modulation of long-term endothelial-barrier integrity is conditional to the cross-talk between Akt and Src signaling. Journal of cellular physiology, 232(10), 2599-2609.
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