cd30 rpmi  (DSMZ)

Journal: Cells

Article Title: Antibody–Drug Conjugate Made of Zoledronic Acid and the Anti-CD30 Brentuximab–Vedotin Exert Anti-Lymphoma and Immunostimulating Effects

doi: 10.3390/cells13100862

Figure Lengend Snippet: Bre–Ved–ZA ADC reactivity. ( A ) Bre–Ved–ZA titration. Upper panels: the HL-cell lines KMH2, L428, L540, HS445, and RPMI6666 were incubated with 20–0.2 µg/mL/10 6 cells of Bre–Ved–ZA ADC (upper histograms, red lines) or Bre–Ved (lower histograms, gray lines), followed by the AlexaFluor647-labeled α-hIg antiserum and FACS analysis. Results expressed as Log far red fluorescence intensity (a.u.) vs. the number of cells. ( B ) HL/CD30 + lymphoblastoid cell lines were incubated with 20–2.0–0.2 µg/mL/10 6 cells of Bre–Ved–ZA ADC (red) or Bre–Ved (gray) as in ( B ). Results expressed as mean fluorescence intensity (MFI-AF647, a.u.).

Article Snippet: KMH2 or L428 (from pleural effusion) and L540 (from bone marrow) classical HL-cell lines (purchased and certified from DSMZ GmbH, Braunschweig, Germany), or the CD30 + lymphoblastoid RPMI6666 and HS445 cell lines (provided and certified by ATCC, Manassas, VA, USA) were incubated with serial dilutions (20–0.02 μg/mL/10 6 cells) of Bre–Ved or Bre–Ved–ZA ADC, for 1 h at 4 °C, followed by the AlexaFluor647 antihuman Ig (AF647-α-hIg) labeling.

Techniques: Titration, Incubation, Labeling, Fluorescence

Journal: Cells

Article Title: Antibody–Drug Conjugate Made of Zoledronic Acid and the Anti-CD30 Brentuximab–Vedotin Exert Anti-Lymphoma and Immunostimulating Effects

doi: 10.3390/cells13100862

Figure Lengend Snippet: HL-cell-line proliferation and apoptosis upon Bre–Ved–ZA or Bre–Ved treatment. ( A ) ATP content, determined using the CellTiter-Glo ® Luminescent Cell Viability Kit, in KMH2, L428, L540, HS445, or RPMI6666 exposed to serial dilution of Bre–Ved or Bre–Ved–ZA (20, 2.0, 0.2 μg/mL) and incubated at 37 °C for 5 days. Luminescence was detected with the VICTORX5 reader and expressed as luminescence arbitrary units (a.u). ( B ) Caspase activation in KMH2 and RPMI6666 cell lines evaluated at 72 h with the luciferase-based Caspase Glo 3/7 3D Assay, expressed as luminescence units (RLU)/5 × 10 4 cells. Results are the mean ± SD of three independent experiments from 4 replicated wells. *** p < 0.001 of Bre–Ved–Za vs. Bre–Ved. ( C ) KMH2 cultured for 72 h with 20 or 2.0 or 0.2 μg/mL Bre–Ved–ZA were labeled with FITC-annexin-V (AV) and propidium iodide (PI) and subjected to FACS analysis. CTR: staining in untreated cells. Apoptotic cells are identified as AV + PI + . ( D ) Confocal microscopy of KMH2 cells treated for 72 h with 2 μg/mL Bre–Ved–ZA upon nuclear staining with Syto16 (left images). Central images: bright field; right images: merged dark and bright fields. Magnification: 20× objective. Bar: 10 µm. ( E ) Confocal microscopy as in ( D ) at higher magnification (40× objective). Bar: 10 µm. Results are representative of three independent experiments. CTR: KMH2 cultured in medium without antibody. The white arrows in ( D , E ) indicate the apoptotic nuclei.

Article Snippet: KMH2 or L428 (from pleural effusion) and L540 (from bone marrow) classical HL-cell lines (purchased and certified from DSMZ GmbH, Braunschweig, Germany), or the CD30 + lymphoblastoid RPMI6666 and HS445 cell lines (provided and certified by ATCC, Manassas, VA, USA) were incubated with serial dilutions (20–0.02 μg/mL/10 6 cells) of Bre–Ved or Bre–Ved–ZA ADC, for 1 h at 4 °C, followed by the AlexaFluor647 antihuman Ig (AF647-α-hIg) labeling.

Techniques: Serial Dilution, Incubation, Activation Assay, Luciferase, Cell Culture, Labeling, Staining, Confocal Microscopy