Journal: Analytical chemistry

Article Title: Rapid Enrichment and Detection of Extracellular Vesicles Enabled by CuS-Enclosed Microgels

doi: 10.1021/acs.analchem.9b04485

Figure Lengend Snippet: Quantification of EV concentrations with the 300-nm CuS-MG in 1× PBS or Serum. Calibration curves in 1×PBS using a) anti-CD63, b) anti-HER2; and in serum using c) anti-CD63 or d) anti-HER2. CL0 = chemiluminescence without EVs. [ABEI] = 0.5 mM, [H2O2] =1 mM, pH=11.

Article Snippet: The biotinylated mouse anti-human ErbB2/HER2 (Recombinant Monoclonal Human IgG 1 Clone Hu5), mouse anti-human CD63 (Clone mem-259), and biotinylated mouse anti-human CD63 (Clone NVG-2) was obtained from R&D systems, Sino Biological, and BioLegend, respectively.

Techniques:

Journal: Analytical chemistry

Article Title: Rapid Enrichment and Detection of Extracellular Vesicles Enabled by CuS-Enclosed Microgels

doi: 10.1021/acs.analchem.9b04485

Figure Lengend Snippet: Detection of cell released EVs using CuS-MG. The cell culture medium was collected from three cell lines: MCF-10A, MDA-MB-231 and SK-BR-3. a) Chemiluminescence resulted from EV detection in the culture media of three cell lines targeting CD63 and anti-HER2; b) PCA plot using the chemiluminescence data shown in a); c) Linear correlation between CD63 quantification results obtained by ELISA and the chemiluminescent signals from CuS-MG in our assay targeting CD63. [ABEI] = 0.5 mM, [H2O2] = 1 mM, pH=11.

Article Snippet: The biotinylated mouse anti-human ErbB2/HER2 (Recombinant Monoclonal Human IgG 1 Clone Hu5), mouse anti-human CD63 (Clone mem-259), and biotinylated mouse anti-human CD63 (Clone NVG-2) was obtained from R&D systems, Sino Biological, and BioLegend, respectively.

Techniques: Cell Culture, Enzyme-linked Immunosorbent Assay

Journal: American Journal of Translational Research

Article Title: Boeravinone B a natural rotenoid exerts anticancer activity via inducing internalization and degradation of inactivated EGFR and ErbB2 in human colon cancer cells

doi:

Figure Lengend Snippet: Effect of Boeravinone B on levels of p-ErbB2, p-ErbB3 and EGFR. A. The human colon cancer HT-29 cells were treated with 0, 1, 3 and 10 µM concentration of Boeravinone B for 24 h followed by western blot analysis for expression of ErbB2, ErbB3, EGFR and transferrin (TfR) B. The HT-29 cells were treated with Boeravinone B (10 µM) for 1, 3, 6, 12 and 24 h. The cell lysates were subjected to western blot analysis along with antibodies specific for ErbB2, p-ErbB2, ErbB3, p-ErbB3, EGFR, p-EGFR and TfR. C. The HT-29 cells were treated with predefined concentrations of Boeravinone B for 3 h. The cell lysates were subjected to western blot analysis using antibodies specific for AKT, pAKT, Erk1/2 and p-Erk1/2. In all the three experiments the blots were compared against loading control α-tubulin.

Article Snippet: Antibodies for AIF, PARP, Erk1/2, caspase-3, phospho-EGFR (Tyr1068), phospho-ErbB3 (Tyr1289), phospho-AKT (Ser473), phospho-Erk1/2 (Thr202/Tyr204), phospho-ErbB2 (Tyr877), Erk1/2, EGFR, GAPDH, a-tubulin, ErbB2, transferring and ErbB3 were obtained from Cell Signaling Tech.

Techniques: Concentration Assay, Western Blot, Expressing, Control

Journal: American Journal of Translational Research

Article Title: Boeravinone B a natural rotenoid exerts anticancer activity via inducing internalization and degradation of inactivated EGFR and ErbB2 in human colon cancer cells

doi:

Figure Lengend Snippet: Effect of Boeravinone B on activation of ErbB2, ErbB3 and EGFR mediated by EGF and HRG. A. The HT-29 cells were pre-treated with Boeravinone B at defined concentrations for 3 h followed by treatment with EGF (50 ng/ml) for 15 min. The whole cell lysates were incubated with antibodies specific for ErbB2, and phospho-ErbB2 and EGFR, phospho-EGFR. B. The human colon cancer HT-29 cells were pre-exposed with 10 µM concentration of Boeravinone B for 3 h followed by treatment of 15 with EGF (50 ng/ml). The lysates were subjected to western blot analysis with antibodies specific for Erk1/2 pErk1/2, AKT and pAKT. C. The HT-29 cells after exposure to Boeravinone B for 3 h followed by treatment with HRG (50 ng/ml) for 15 min followed by stimulation with HRG (50 ng/ml). Western blot analysis was done for lysate along with antibodies specific for ErbB2, p-ErbB2, ErbB3 and p-ErbB3. D. The HT-29 cells were pre-exposed with 10 µM concentration of Boeravinone B for 3 h followed by treatment with HRG (50 ng/ml) for 15 min. The lysates were subjected to western blot analysis with antibodies specific for Erk1/2, pErk1/2, AKT and p-AKT.

Article Snippet: Antibodies for AIF, PARP, Erk1/2, caspase-3, phospho-EGFR (Tyr1068), phospho-ErbB3 (Tyr1289), phospho-AKT (Ser473), phospho-Erk1/2 (Thr202/Tyr204), phospho-ErbB2 (Tyr877), Erk1/2, EGFR, GAPDH, a-tubulin, ErbB2, transferring and ErbB3 were obtained from Cell Signaling Tech.

Techniques: Activation Assay, Incubation, Concentration Assay, Western Blot

Journal: American Journal of Translational Research

Article Title: Boeravinone B a natural rotenoid exerts anticancer activity via inducing internalization and degradation of inactivated EGFR and ErbB2 in human colon cancer cells

doi:

Figure Lengend Snippet: Boeravinone B leads to internalization of ErbB2 and EGFR in human colon cancer HT-29 cells. A. The HT-29 cells were treated with 10 µM concentration of Boeravinone B for 24 h along with Lactacystin (5 µM) and Chloroquine (30 µM) followed by western blot analysis of lysates along with ErbB2 and EGFR antibodies. B. Western blot analysis for conforming internalization of ErbB2, EGFR and TfR by surface biotinylation assay in HT-29 cells treated with Boeravinone B, control and positive control (EGF treated for 0.5 h). C. The colon cancer HT-29 cells were treated with 10 µM of Boeravinone B for 0, 3 and 24 h, after fixation and incubating cells with anti-EGFR or anti-ErbB2 antibodies followed by subsequent exposure to Alexa 546-conjugated anti-rabbit secondary followed by microscopic analysis.

Article Snippet: Antibodies for AIF, PARP, Erk1/2, caspase-3, phospho-EGFR (Tyr1068), phospho-ErbB3 (Tyr1289), phospho-AKT (Ser473), phospho-Erk1/2 (Thr202/Tyr204), phospho-ErbB2 (Tyr877), Erk1/2, EGFR, GAPDH, a-tubulin, ErbB2, transferring and ErbB3 were obtained from Cell Signaling Tech.

Techniques: Concentration Assay, Western Blot, Surface Biotinylation Assay, Control, Positive Control

Journal: Cancers

Article Title: Early Detection and Dynamic Changes of Circulating Tumor Cells in Transgenic NeuN Transgenic (NTTg) Mice with Spontaneous Breast Tumor Development

doi: 10.3390/cancers13133294

Figure Lengend Snippet: Surface modification and the non-fouling effect monitored by QCM-D. The kinetics of the stepwise formation of the surface coating, monitored by normalized frequency F using QCM-D for coating with mEpCAM antigens. Lipid vesicles containing 4 mol% of b-PE were introduced at point (I) and absorbed and ruptured to form bSLB on a silicon dioxide chip. NeutrAvidin (NA) solution was injected at point (II) and incubated for 20 h for specific binding on SLB, forming NA-bSLB. Biotinylated anti-mEpCAM IgG solution was added at point (III) to conjugate anti-mEpCAM to the NA-bSLB layer. Then, recombinant mEpCAM was injected at point (IV) to determine the IgG-antigen binding ratio. The pH 7.2 PBS buffer rinse, denoted as *, was performed at least once after each step.

Article Snippet: Afterward, the procedures of glass surface modification were modified from the previous study by coating with biotinylated anti-mouse-EpCAM IgG (13-5791-82, eBioscience, Thermo Fisher Scientific, Waltham, MA, USA) (mCMx-EpCAM chip) or biotinylated anti-rat-HER2 IgG (610161, BD Bioscience, San Jose, CA) (mCMx-HER2 chip).

Techniques: Modification, QCM-D, Injection, Incubation, Binding Assay, Recombinant

Journal: Cancer Genomics & Proteomics

Article Title: Serum Exosomes Expressing CD9, CD63 and HER2 From Breast-Cancer Patients Decreased After Surgery of the Primary Tumor: A Potential Biomarker of Tumor Burden

doi: 10.21873/cgp.20474

Figure Lengend Snippet: Pre- and post-operative CD9 and HER2 double-positive serum exosome quantification. CD9 and HER2 double-positive exosomes in the serum of breast-cancer patients (n=5) were quantitated pre- and post-operatively. Data were statistically analyzed using the Mann-Whitney U-test (p<0.05).

Article Snippet: Biotin labeling of HER2 antibody (sc-33684, Santa Cruz Biotechnology, Dallas, TX, USA) was performed by binding to avidin-conjugated beads using the Biotin labeling kit-NH 2 (Dojindo Laboratories, Kumamoto, Japan).

Techniques: MANN-WHITNEY