Journal: mAbs

Article Title: Generation and in vivo characterization of a novel high-affinity human antibody targeting carcinoembryonic antigen

doi: 10.1080/19420862.2023.2217964

Figure Lengend Snippet: In vitro characterization of new anti-CEA antibodies. (a) SPR sensorgram of G9, F7, and F4 in a scFv format. K D values were calculated to be 640 nM, 50 nM, and 7.7 nM, respectively. (b) Flow cytometry analysis with the new antibodies in IgG format on CEA-expressing CT26 cells and on CEA-negative CT26 wild-type cells. (c) Immunofluorescence staining with IgG formats on the human colon adenocarcinoma xenograft LS174T. Anti-CEA antibodies were detected in green. Blood vessels were detected by CD31 staining (red). 20× magnification, scale bars = 100 μm.

Article Snippet: Immunofluorescence analysis was performed on LS174T and HT-29 xenografts harvested from mice, on patient-derived colon cancer samples, and on a human tissue microarray (Amsbio, T6235700–5).

Techniques: In Vitro, Flow Cytometry, Expressing, Immunofluorescence, Staining

Journal: mAbs

Article Title: Generation and in vivo characterization of a novel high-affinity human antibody targeting carcinoembryonic antigen

doi: 10.1080/19420862.2023.2217964

Figure Lengend Snippet: Ex vivo immunofluorescence-based biodistribution analysis. Immunofluorescence analysis assessed tumor targeting of new anti-CEA antibodies in IgG format. Two hundred micrograms of IgG-FITC were injected intravenously into LS174T-bearing mice. Tumors were excised 24 hours after injection. IgG-FITC was detected in green; blood vessels were detected through CD31 staining (red). 20× magnification, scale bars = 100 μm.

Article Snippet: Immunofluorescence analysis was performed on LS174T and HT-29 xenografts harvested from mice, on patient-derived colon cancer samples, and on a human tissue microarray (Amsbio, T6235700–5).

Techniques: Ex Vivo, Immunofluorescence, Injection, Staining

Journal: mAbs

Article Title: Generation and in vivo characterization of a novel high-affinity human antibody targeting carcinoembryonic antigen

doi: 10.1080/19420862.2023.2217964

Figure Lengend Snippet: Quantitative biodistribution with radiolabeled anti-CEA antibodies in diabody format. Quantitative biodistribution analysis of radio iodinated anti-CEA diabodies in BALB/c nude mice bearing subcutaneous LS174T colon adenocarcinomas. Organs were harvested 24 hours after intravenous injection, and radioactivity was quantified. Results are shown as the percentage of injected dose per gram (ID/g (%)). Error bars = SEM; n = 4.

Article Snippet: Immunofluorescence analysis was performed on LS174T and HT-29 xenografts harvested from mice, on patient-derived colon cancer samples, and on a human tissue microarray (Amsbio, T6235700–5).

Techniques: Injection, Radioactivity

Journal: International Journal of Molecular Sciences

Article Title: Novel Lipid Nanocomplex Co-Carrying Bcl2 siRNA and Quantum Dots for EGF Receptor-Targeted Anti-Cancer Theranosis

doi: 10.3390/ijms25116246

Figure Lengend Snippet: Tumor-targeted siRNA delivery of QDM and immuno-QDM. Protein expression of MDA-MB-453 (EGFR negative) and LS174T (EGFR positive) cells, and the target tumor cell binding and cellular uptake assay of Chol-siRNA, QDM, and immuno-QDM ( A – C ) were evaluated. All particles were incubated with the target cells (LS174T) for 1 h and 8 h at 37 °C and analyzed with a confocal microscope for evaluate siRNA delivery and endosomal escape analysis of QDM and immuno-QDM ( D ) (X400).

Article Snippet: The LS174T (human colorectal adenocarcinoma, #10188) cell line was purchased from the Korean Cell Line Bank (Seoul, Republic of Korea).

Techniques: Expressing, Binding Assay, Incubation, Microscopy

Journal: International Journal of Molecular Sciences

Article Title: Novel Lipid Nanocomplex Co-Carrying Bcl2 siRNA and Quantum Dots for EGF Receptor-Targeted Anti-Cancer Theranosis

doi: 10.3390/ijms25116246

Figure Lengend Snippet: Knockdown of target gene and in vitro anti-cancer efficacy of QDM/siBcl2 and iQDM/siBcl2. ( A , B ) The bcl2 gene knockdown was detected with PCR and Western blot (25, 50 pm). ( C ) MTT assay results of LS174T and MDA-MB-453 cells after the treatment of siRNA, QDM/siBcl2, and iQDM/siBcl2.

Article Snippet: The LS174T (human colorectal adenocarcinoma, #10188) cell line was purchased from the Korean Cell Line Bank (Seoul, Republic of Korea).

Techniques: Knockdown, In Vitro, Western Blot, MTT Assay

Journal: Advanced biosystems

Article Title: Analyzing Mechanisms of Metastatic Cancer Cell Adhesive Phenotype Leveraging Preparative Adhesion Chromatography Microfluidic

doi: 10.1002/adbi.201800328

Figure Lengend Snippet: (A) Elution profiles of LS174T metastatic colon carcinoma cell populations when perfused over E-selectin, demonstrating the early elution of free flow cells followed by subsequent elution of adherent cells. (B) Hemodynamic microenvironment-mimicking, parallel-plate microfluidic fabrication, into which (C, bottom) metastatic cancer cells are infused into the E-selectin functionalized channel, visualized via an integrated high speed videomicroscopy, and collected into collection tubes based on channel residence time. (D-H) Continuous perfusion experiments, (I-M) Static perfusion experiments. (D,I) Channel functionalization schematic and perfusion workflow diagram. (E) Distance of individual perfused cells from the channel bottom calculated based on measured individual cell velocity and size decreases from the inlet (channel position 1) until reaching channel position 3 after which it is uniform throughout the remaining channel length. Data represent mean ± s.e.m. (F,K) Representative relative frequency distribution of elution time of the infused cell pulse with the inset showing the mean time at which 95% of the cell pulse has eluted ± s.e.m. (G,L) Number of interacting cells along the length of the channel during the sorting phase. Data represent mean ± s.e.m. (H,M) Number of interacting cells along the width of the channel (black) and average velocity of rolling cells along the width of the channel (gray). Data represent mean ± s.e.m. (J) Location of cell pulse during static phase of Static perfusion experiments (E-H,J-M) Each point represents the mean ± s.e.m. of n≥5 independently run experiments. Cell pulse of LS174T colon carcinoma cells perfused at 1 dyn cm−2 over (E,G,I,K) 2.5 ug mL−1 E-selectin or (F,J) unfunctionalized channels. * p < 0.05 ** p<0.01 compared to unfunctionalized channel (no interacting cells) by one-sample t-test.

Article Snippet: In Vivo Metastasis Model: 10 4 LS174T cells, either from the unsorted parental population or fractionated using the adhesion chromatography system, were suspended in 100 μL of sterile saline and each fraction was injected intravenously via the tail vein into six week old female NSG mice under isoflurane anesthesia (Jackson Labs).

Techniques:

Journal: Advanced biosystems

Article Title: Analyzing Mechanisms of Metastatic Cancer Cell Adhesive Phenotype Leveraging Preparative Adhesion Chromatography Microfluidic

doi: 10.1002/adbi.201800328

Figure Lengend Snippet: (A) Percent of LS174T cells collected in either the free flow or adherent fraction of the parent (unsorted) population or reperfused cell subpopulations either immediately after perfusion or after 2 passages. Each bar represents the mean ± s.e.m. of n≥3 individually run experiments. (B) Quantification of sLex expression of cells in each collected cell fraction. (C,D,H,I) mRNA expression heatmaps of (C) all analyzed mRNA, (D) genes regulating CEA, CD44, and sLex presentation, (H) actin-cytoskeleton structure, and (I) cell cycle stage. US0/FF0/AD0: unsorted/free flow/adherent, flash frozen at 0 hr; US48/FF84/AD48: unsorted/free flow/adherent, flash frozen at 48 hr. (E-G) Histograms of log2 fold change at 0 hr of genes regulating CEA, CD44, and sLex presentation of (E) free flow compared to adherent, (F) free flow compared to unsorted, and (G) adherent compared to unsorted. (A-C) Perfusion and separation of an LS174T cell pulse of 250,000 cells was over 2.5 μg mL−1 E-selectin at 1 dyn cm−2. * p < 0.05 *** p < 0.001 **** p< 0.0001 by (A) One way ANOVA with post-hoc t-test or (B) t-test.

Article Snippet: In Vivo Metastasis Model: 10 4 LS174T cells, either from the unsorted parental population or fractionated using the adhesion chromatography system, were suspended in 100 μL of sterile saline and each fraction was injected intravenously via the tail vein into six week old female NSG mice under isoflurane anesthesia (Jackson Labs).

Techniques: Expressing

Journal: Advanced biosystems

Article Title: Analyzing Mechanisms of Metastatic Cancer Cell Adhesive Phenotype Leveraging Preparative Adhesion Chromatography Microfluidic

doi: 10.1002/adbi.201800328

Figure Lengend Snippet: (A) Representative bright field and chromogenic images of NSG mouse whole lobes and lung sections and 7 d post intravenous (i.v.) infusion of parent (unsorted) or sorted free flow or adherent LS174T cells. Arrows indicate metastatic tumors. Scale bars, 100 μm or 1 mm in regions of interest and whole lobe images, respectively. (B) Lung metastasis count in lungs dissected from NSG mice either 7 or 14 d post i.v. infusion of collected LS174T free flow cells or adherent cells. (C) Count of lung metastases in either the free flow group or the adherent group normalized to the mean number of lung metastases per NSG mouse in the parent (unsorted) group 7 d post i.v. infusion. (D) Quantification of hematoxylin and eosin staining for each group. (A-D) Perfusion and separation of an LS174T cell pulse of 250,000 cells was over 2.5 μg mL−1 E-selectin at 1 dyn cm−2. NSG mice were injected i.v. with 105 LS174T cells. (B-C) Each point represent an individual mouse lung. Bars represent mean ± s.e.m. of n≥5 individual mice. (B) * p < 0.05 by two sample t-test. (C) * p < 0.05 by t-test Ho: μ= 1 (D) Bars represent mean ± SEM of independent samples. Analyzed with a one-way ANOVA with post hoc t-test with Bonferonni corrections for multiple comparisons * p < 0.05 ** p < 0.01

Article Snippet: In Vivo Metastasis Model: 10 4 LS174T cells, either from the unsorted parental population or fractionated using the adhesion chromatography system, were suspended in 100 μL of sterile saline and each fraction was injected intravenously via the tail vein into six week old female NSG mice under isoflurane anesthesia (Jackson Labs).

Techniques: Staining, Injection

Journal: Advanced biosystems

Article Title: Analyzing Mechanisms of Metastatic Cancer Cell Adhesive Phenotype Leveraging Preparative Adhesion Chromatography Microfluidic

doi: 10.1002/adbi.201800328

Figure Lengend Snippet: (A,D,G) Representative flow cytometric histograms of sLex (HECA452), CD44, or CEA expression. (B,E,H) sLex (HECA452), CD44, or CEA expression normalized to the parent population between cells in the collected free flow fraction and adherent fraction. Each point represents the mean fluorescent intensity of analyzed cells ± s.e.m. (C,F,I) Percent of cells in each fraction that are in the top 10% or bottom 10% of sLex (HECA), CD44, or CEA expressing cells. Each bar represents the mean ± s.e.m. of n≥3 individually run experiments. (J) Representative flow cytometry gating and analysis strategy for (K,L) sLex (HECA452) expression levels normalized to the unsorted parent LS174T cell population in either high (top 10%) or low (bottom 10%) expressing subpopulations of (K) CD44 or (L) CEA. Each bar represents the mean ± s.e.m. of n≥3 individually run experiments. Perfusion and separation of an LS174T cell pulse of 250,000 cells was over 2.5 μg mL−1 E-selectin at 1 dyn cm−2. * p < 0.05 ** p < 0.01 *** p, 0.001 by paired t-test.

Article Snippet: In Vivo Metastasis Model: 10 4 LS174T cells, either from the unsorted parental population or fractionated using the adhesion chromatography system, were suspended in 100 μL of sterile saline and each fraction was injected intravenously via the tail vein into six week old female NSG mice under isoflurane anesthesia (Jackson Labs).

Techniques: Expressing, Flow Cytometry

Journal: Advanced biosystems

Article Title: Analyzing Mechanisms of Metastatic Cancer Cell Adhesive Phenotype Leveraging Preparative Adhesion Chromatography Microfluidic

doi: 10.1002/adbi.201800328

Figure Lengend Snippet: (A-C) Representative flow cytometric histograms of sLex (HECA452), CD44, or CEA expression indicating gating for the top and bottom 10% percent of cells expressing the respective ligand and the CD24 MFI in the highest versus lowest 10% of HECA, CD44, or CEA expressing cells. (D) Average mean fluorescent intensity of cells obtained from the free flow and adherent fractions in continuous flow. (E) Percentage of cells in the top or bottom 10% of CD24 expressing cells after continuous flow. (D,E) Each bar represents the mean ± s.e.m. of n≥3 individually run experiments. (F-H) Comparisons between frequencies of cells in low versus high selectin ligand expression derived from costaining. (A-C, F-H) Each bar represents the mean ± s.e.m. of n≥3 individually run experiments. Perfusion and separation of an LS174T cell pulse of 250,000 cells was over 2.5 μg mL−1 E-selectin at 1 dyn cm−2. * p < 0.05 ** p < 0.01 by paired t-test.

Article Snippet: In Vivo Metastasis Model: 10 4 LS174T cells, either from the unsorted parental population or fractionated using the adhesion chromatography system, were suspended in 100 μL of sterile saline and each fraction was injected intravenously via the tail vein into six week old female NSG mice under isoflurane anesthesia (Jackson Labs).

Techniques: Expressing, Derivative Assay