TNF-α was purchased from PeproTech (Rocky Hill, NJ, USA). Primary antibodies specific for α-tubulin (T5168) and vimentin (V6630) were purchased from Sigma-Aldrich (St. Louis, MO, USA). Primary antibodies specific for VCAM-1 (ab134047), MGMT (ab108630), and β-catenin (ab16051) were purchased from Abcam (Cambridge, UK). Primary antibodies specific for AKT1/2/3 (sc-8312), p-AKT1/2/3 (Ser473, sc-7985-R), connexins 43 (sc-271837), eIF2α (sc-133132), and N-cadherin (sc-7939) were purchased from Santa Cruz Biotechnology (Santa Cruz, CA, USA). Primary antibodies specific for EGFR (4267), p-Cx43 (Ser368, 3511), and p-mTOR (Ser2448, 2971) were purchased from Cell Signaling Technology (Danvers, MA, USA). Primary antibodies specific for p-GSK3β (Tyr216, 44604G) was purchased from Invitrogen (Carlsbad, CA, USA).
N cadherin sc 7939
Manufactured by Santa Cruz Biotechnology
Sourced in United States
N-cadherin sc-7939 is a primary antibody that recognizes the N-cadherin protein. N-cadherin is a cell adhesion molecule involved in cell-cell interactions. This product can be used for research applications that require the detection of N-cadherin.
Automatically generated - may contain errors
Lab products found in correlation
Vimentin sc 6260, by Santa Cruz Biotechnology (3 mentions)
E cadherin sc 8426, by Santa Cruz Biotechnology (3 mentions)
Pierce ecl western blotting substrate, by Thermo Fisher Scientific (3 mentions)
Snail ab167609, by Abcam (2 mentions)
Twist ab50887, by Abcam (2 mentions)
Ab16051, by Abcam (1 mentions)
Ab108630, by Abcam (1 mentions)
V6630, by Merck Group (1 mentions)
Ab134047, by Abcam (1 mentions)
Sc 8312, by Santa Cruz Biotechnology (1 mentions)
Eif2α sc 133132, by Santa Cruz Biotechnology (1 mentions)
T5168, by Merck Group (1 mentions)
Tnf α, by Thermo Fisher Scientific (1 mentions)
Sodium dodecyl sulfate (sds), by Thermo Fisher Scientific (1 mentions)
Bromophenol blue, by Merck Group (1 mentions)
Glycerol, by Avantor (1 mentions)
Image studio lite software, by LI COR (1 mentions)
Horseradish peroxide hrp conjugated secondary antibody, by Santa Cruz Biotechnology (1 mentions)
Coomassie plus protein assay, by Thermo Fisher Scientific (1 mentions)
Cellytic m cell lysis reagent, by Merck Group (1 mentions)
6 protocols using n cadherin sc 7939
1
Protein Expression Analysis Protocol
2
Mesensphere Protein Quantification and Cadherin Analysis
3
Western Blot Analysis of EMT Markers
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Cells were lysed on ice in RIPA (radio-immunoprecipitation assay) buffer with 1 mM PMSF (phenylmethylsulfonyl fluoride). From the cell lysate, 40 μg of total protein was loaded into the wells of the SDS-PAGE (sodium dodecyl sulfate polyacrylamide gel elec trophoresis) gel, along with a molecular weight marker. Electrophoresis was carried out for 1–2 h at 100 V, followed by transfer to PVDF (polyvinyl difluoride) membranes, which were then blocked with 5% BSA (bovine serum albumin). The membranes were then incubated overnight at 4°C with primary antibodies (E-cadherin sc-8426, N-cadherin sc-7939, vimentin sc-6260, Santa Cruz; fibulin-4 ab125073, Snail ab167609, Slug ab27568, Twist ab50887, PI3K ab86714, p-PI3K ab182651, AKT ab8805, P-AKT 38449, mTOR ab32028, p-mTOR ab109268; Abcam) at working dilutions of 1:1000. After washing the membranes thrice with TBST for 5 min each, the membranes were incubated with conjugated secondary antibody diluted to 1:1000, at room temperature for 1 h. Blots were developed using the enhanced chemiluminescence method (Pierce™ ECL Western Blotting Substrate; Thermo Fisher Scientific, Inc.).
4
Western Blot Analysis of EMT Markers
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Using radioimmunoprecipitation assay buffer supplemented with 1 mM phenylmethylsulfonyl fluoride, cells were lysed on ice. Protein samples (40 μg/lane) were subjected to sodium dodecyl sulfate polyacrylamide gel electrophoresis, transferred to polyvinyl difluoride membranes and blocked with 5% bovine serum albumin. These membranes were then cultured with primary antibodies (E-cadherin sc-8426, N-cadherin sc-7939, vimentin sc-6260, Santa Cruz; HMGB1 ab18256, Snail ab167609, Twist ab50887, Abcam) with working dilutions 1:1,000 at 4°C overnight. The next day, these membranes were incubated with secondary antibody with working dilutions 1:2,000 at room temperature for 1 h, and blots were developed using the enhanced chemiluminescence method (Pierce™ ECL Western Blotting Substrate; Thermo Fisher Scientific, Inc.).
5
Western Blot Analysis of EMT Markers
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Cells were lysed on ice in RIPA (radio-immunoprecipitation assay) buffer containing 1 mM phenylmethylsulfonyl fluoride (PMSF). Samples (40-μg protein per lane) were separated by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE), transferred to polyvinyl difluoride (PVDF) membranes, and blocked with 5% bovine serum albumin (BSA). Membranes were then incubated with primary antibodies (E-cadherin sc-8426, N-cadherin sc-7939, Vimentin sc-6260, Santa Cruz; fibulin-4 ab125073, Snail ab167609, Slug ab27568, Twist ab50887, β-catenin ab32572, C-myc ab32072, Cyclin D1 ab134175, Notch1 intracellular domain (NICD) ab83232, HES 1 ab71559, HEY L ab154077, Abcam) at working dilutions of 1:1000 at 4°C overnight. On the next day, membranes were incubated with secondary antibody for 1 hour at room temperature, and blots were developed using the enhanced chemiluminescence method (Pierce™ ECL Western Blotting Substrate; Thermo Fisher Scientific, Inc.).
6
PBrP Modulation of TGF-β Signaling
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To examine the effect of PBrP on TGF-β-induced Smad2/3 phosphorylation, Mv1Lu and A549 cells were incubated with PBrP at various concentrations or for various periods and then stimulated with 100 pM of TGF-β for 30 min. To monitor EMT protein expression, the cells were pretreated with different PBrP concentrations for 2 h and then stimulated with TGF-β for 48 h. The cells were lysed with a lysis buffer and standardised using a bicinchoninic acid (BCA) protein assay (Pierce™ BCA Protein Assay Kit 23225). Fifty microgram of this protein was applied to SDS-PAGE and electrotransferred to the polyvinylidene difluoride (PVDF) membrane. Polyclonal antibodies against TβRI (sc-398), TβRII (sc-400), EGFR (sc-373746), β-actin (sc-47778), flotilin-1 (sc-25506), fibronectin (sc-9068), N-cadherin (sc-7939), Lamin B (sc-6216) and vimentin (sc-7557) were obtained from Santa Cruz (Dallas, TX). Rabbit Polyclonal antibodies against pSmad2/3 (#8828), Smad2/3 (#8685), PAI-1 (#11907), Flag tag (#8146), Lamp-1 (sc-9091) and MyoVa (#3402) were purchased from Cell Signalling (Boston, MA). Secondary antibodies conjugated with horseradish peroxidase (Millipore, Darmstadt, Germany) and an enhanced chemiluminescence (ECL) kit (Perkin-Elmer Life Sciences, Waltham, MA, USA) were used to develop immunoblots.
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