Journal: Cancers

Article Title: Landscape of 4D Cell Interaction in Hodgkin and Non-Hodgkin Lymphomas

doi: 10.3390/cancers13205208

Figure Lengend Snippet: Number of cells analyzed in the different cases.

Article Snippet: Live vibratome sections were stained for 15 min at 37 °C with the following antibodies: Alexa Fluor 647–anti-human PD-1 (clone EH12.2H7; BioLegend, San Diego, CA, USA), Alexa Fluor 647–anti-human CD30 (clone CD30/412, Novus Biologicals, Centennial, CO, USA), Alexa Fluor 488–anti-human PD-1 (clone EH12.2H7; BioLegend), Alexa Fluor 647–anti-human CD3 (clone UCHT1; BD Biosciences), Brilliant Violet 421–anti-human CD3 (clone UCHT1; BioLegend), Alexa Fluor 488–anti-human CD19 (clone HIB 19; BioLegend), Pacific Blue–anti-human CD20 (clone 2H7; BioLegend), Alexa Fluor 555-anti-human CD20 (polyclonal rabbit antibody; Bioss Antibodies, Woburn, MA, USA), and Alexa Fluor 555-anti-human CD19 (polyclonal rabbit antibody; Bioss Antibodies).

Techniques:

Journal: Cancers

Article Title: Landscape of 4D Cell Interaction in Hodgkin and Non-Hodgkin Lymphomas

doi: 10.3390/cancers13205208

Figure Lengend Snippet: Velocity of different cell types under reactive and neoplastic conditions. ( a ). Velocity of PD1-positive T cells, CD20-positive B cells and CD30-positive cells in all cases studied (*** p < 0.001, one-way-ANOVA with Bonferroni’s post-test for multiple comparisons). Each dot represents the mean from one movie. ( b ). Velocity of CD20-positive B cells in lymphadenitis and malignant lymphomas. The velocity of bulk CD20-positive B cells in CD20+ lymphomas largely represents the velocity of tumor cells, whereas the velocity in CD20- lymphomas corresponds to reactive bystander B cells. (** p = 0.0042, unpaired t -test.) Each dot represents the mean from one movie. ( c ). Velocity of CD20-positive B cells listed according to different lymphoma entities. No significant differences were observed. Each dot represents the mean from one movie. Lymphadenitis: 4 cases, AITL, DLBCL and MCL: one case, MGZL and NLPHL: two cases, cHL: three cases. ( d ). Velocity of PD1-positive T cells grouped according to diagnosis of lymphadenitis, mature B-cell lymphomas and Hodgkin lymphomas. Each dot represents the mean from one movie. * p < 0.05, ** p < 0.01, Kruskal–Wallis test with Dunn’s post-test for multiple comparisons. ( e ). Velocity of PD1-positive T cells according to lymphoma entities. ** p < 0.01, *** p < 0.001, Kruskal–Wallis-Test with Dunn´s post-test for multiple comparisons. Each dot represents the mean from one movie. Lymphadenitis: 5 cases, AITL and MCL: one case, MGZL, DLBCL, FL and NLPHL: two cases, cHL: three cases. ( f ). Velocity of CD30-positive lymphoma cells compared with CD30-positive reactive bystander cells in CD30-negative lymphomas and lymphadenitis (* p < 0.05, unpaired t -test). Each dot represents the mean from one movie.

Article Snippet: Live vibratome sections were stained for 15 min at 37 °C with the following antibodies: Alexa Fluor 647–anti-human PD-1 (clone EH12.2H7; BioLegend, San Diego, CA, USA), Alexa Fluor 647–anti-human CD30 (clone CD30/412, Novus Biologicals, Centennial, CO, USA), Alexa Fluor 488–anti-human PD-1 (clone EH12.2H7; BioLegend), Alexa Fluor 647–anti-human CD3 (clone UCHT1; BD Biosciences), Brilliant Violet 421–anti-human CD3 (clone UCHT1; BioLegend), Alexa Fluor 488–anti-human CD19 (clone HIB 19; BioLegend), Pacific Blue–anti-human CD20 (clone 2H7; BioLegend), Alexa Fluor 555-anti-human CD20 (polyclonal rabbit antibody; Bioss Antibodies, Woburn, MA, USA), and Alexa Fluor 555-anti-human CD19 (polyclonal rabbit antibody; Bioss Antibodies).

Techniques: Biomarker Discovery

Journal: Cancers

Article Title: Landscape of 4D Cell Interaction in Hodgkin and Non-Hodgkin Lymphomas

doi: 10.3390/cancers13205208

Figure Lengend Snippet: Cell tracks according to different lymphoma types superimposed on fluorescence images. ( a – c ) Tracks of the tumor cells of mantle cell lymphoma ( a ), NLPHL ( b ) and MGZL ( c ) in CD20- or CD30-immunostaining, as indicated. Color of tracks corresponds to the observed velocity as indicated in the scale. ( d – f ) Tracks of the bystander T cells of the same cases: mantle cell lymphoma ( d ), NLPHL ( e ) and MGZL ( f ) in PD1-staining. Color of tracks corresponds to the observed velocity as indicated in the scale. ( g ) HE staining (200x) of the mantle cell lymphoma from ( a , d ). ( h ) HE staining (200x) of the NLPHL from ( b , e ). ( i ) HE staining (200×) of the MGZL from ( c , f ).

Article Snippet: Live vibratome sections were stained for 15 min at 37 °C with the following antibodies: Alexa Fluor 647–anti-human PD-1 (clone EH12.2H7; BioLegend, San Diego, CA, USA), Alexa Fluor 647–anti-human CD30 (clone CD30/412, Novus Biologicals, Centennial, CO, USA), Alexa Fluor 488–anti-human PD-1 (clone EH12.2H7; BioLegend), Alexa Fluor 647–anti-human CD3 (clone UCHT1; BD Biosciences), Brilliant Violet 421–anti-human CD3 (clone UCHT1; BioLegend), Alexa Fluor 488–anti-human CD19 (clone HIB 19; BioLegend), Pacific Blue–anti-human CD20 (clone 2H7; BioLegend), Alexa Fluor 555-anti-human CD20 (polyclonal rabbit antibody; Bioss Antibodies, Woburn, MA, USA), and Alexa Fluor 555-anti-human CD19 (polyclonal rabbit antibody; Bioss Antibodies).

Techniques: Fluorescence, Immunostaining, Staining

Journal: Cancers

Article Title: Landscape of 4D Cell Interaction in Hodgkin and Non-Hodgkin Lymphomas

doi: 10.3390/cancers13205208

Figure Lengend Snippet: Cell–cell contacts between tumor and bystander cells. ( a ) Number of cell–cell contacts per movie between tumor cells (CD20 or CD30 as indicated) and PD1-positive T cells (for AITL PD1-positive tumor cells and CD20-positive B cells). Each dot represents the mean from one movie. Lymphadenitis: 4 cases, AITL, DLBCL, NLPHL and MCL: one case, MGZL: two cases, cHL: three cases. ( b ) Duration of cell–cell contacts between tumor (CD20 or CD30 as indicated) and PD1-positive T cells (for AITL PD1-positive tumor cells and CD20-positive B cells, * p < 0.05, Kruskal–Wallis test with Dunn’s post-test for multiple comparisons). Each dot represents the mean from one movie. Lymphadenitis: 4 cases, AITL, DLBCL, NLPHL and MCL: one case, MGZL: two cases, cHL: three cases.

Article Snippet: Live vibratome sections were stained for 15 min at 37 °C with the following antibodies: Alexa Fluor 647–anti-human PD-1 (clone EH12.2H7; BioLegend, San Diego, CA, USA), Alexa Fluor 647–anti-human CD30 (clone CD30/412, Novus Biologicals, Centennial, CO, USA), Alexa Fluor 488–anti-human PD-1 (clone EH12.2H7; BioLegend), Alexa Fluor 647–anti-human CD3 (clone UCHT1; BD Biosciences), Brilliant Violet 421–anti-human CD3 (clone UCHT1; BioLegend), Alexa Fluor 488–anti-human CD19 (clone HIB 19; BioLegend), Pacific Blue–anti-human CD20 (clone 2H7; BioLegend), Alexa Fluor 555-anti-human CD20 (polyclonal rabbit antibody; Bioss Antibodies, Woburn, MA, USA), and Alexa Fluor 555-anti-human CD19 (polyclonal rabbit antibody; Bioss Antibodies).

Techniques:

Journal: Aesthetic surgery journal

Article Title: IL-9 Is a Biomarker of BIA-ALCL Detected Rapidly by Lateral Flow Assay.

doi: 10.1093/asj/sjae137

Figure Lengend Snippet: Figure 3. CD30 lateral flow assay. (A) CD30 standards were diluted from 0 to 10 ng/mL in a benign seroma. (B) Correlation of CD30 concentration with TL/CL ratio (r2 = 0.72, y = 0.05x + 0.7, P = .02). (C) Comparison of CD30 in benign and ALCL seromas (benign, 0.198 ± 0.036 [n = 15] vs ALCL, 0.779 ± 0.083 [n = 11]; P < .0001, t-test). CL, control line; TL, test line.

Article Snippet: Similarly, CD30 LFA strips were made by striping mouse anti-CD30 antibody (R&D Systems, catalog number MAB2291).

Techniques: Lateral Flow Assay, Concentration Assay, Comparison, Control

Journal: Cancer research

Article Title: CD30 is involved in inhibition of T-cell proliferation by Hodgkin's Reed-Sternberg cells.

doi: 10.1158/0008-5472.can-03-1337

Figure Lengend Snippet: Fig. 2. A, confocal microscopy of CD30-ex- pressing CHO cells stained with phycoerythrin- conjugated anti-CD30. B, detection of CD30 by Western blot. The estimated Mr of the EGFP- tagged CD30 is 140,000, that of CD30 is 110,000. Lane 1, CD30 CHO cells; Lane 2, HDLM cells; Lane 3, CD30/EGFP CHO cells; Lane 4, CD30 CHO cells; Lane 5, CD30-Fc fusion pro- tein in reduced condition (estimated Mr, 80,000); and Lane 6, fusion protein in nonreduced condition (Mr 160,000). C, CHO cells with or without CD30 expression were examined for the inhibitory effect on T-cell proliferation. CD30CHO cells, but not EGFPCHO cells, were able to inhibit the tritium uptake of anti-CD3-stimulated T cells. On the other hand, anti-CD30 treatment abolished the inhibitory effect of CD30CHO cells on T-cell proliferation. Similar effects were obtained with mitomycin-C- treated or paraformaldehyde-fixed CD30CHO cells (not shown). CHOEGFP, EGFP CHO cells; CHOCD30, CD30 CHO cells; 30-CHOCD30, anti-CD30-treated CD30 CHO cells; 40- CHOCD30, anti-CD40-treated CD30 CHO cells.

Article Snippet: Immunostaining was performed with goat polyclonal primary antibodies specific for human CD30 (R&D, Minneapolis, MN), followed by incubation with rabbit polyclonal antigoat IgG antibody conjugated to horseradish peroxidase (Santa Cruz Biotechnology, Santa Cruz, CA).

Techniques: Confocal Microscopy, Staining, Western Blot, Expressing

Journal: Cancer research

Article Title: CD30 is involved in inhibition of T-cell proliferation by Hodgkin's Reed-Sternberg cells.

doi: 10.1158/0008-5472.can-03-1337

Figure Lengend Snippet: Fig. 1. H-RS cells inhibit T-cell proliferation, and the inhibition is CD30 dependent. A, T cells and accessory cells (each 1 105 cells/well) were cocultured with soluble anti-CD3. Cells were analyzed for the expression of CD30 and CD153 at the start of and 24 h after coculture. Anti-CD3-treated T cells did not express CD30 and expressed CD153 after activation. B, H-RS cells (only KM-H2 cells are shown) expressed CD30 but no or little CD153. C, anti-CD3-treated T cells, accessory cells, and mitomycin C-pretreated H-RS cells (e.g., KM-H2, HDLM, and L428), or U937 cells, in different ratios to T-cell number (H/T ratio), were cocultured with anti-CD3 at 0.5 g/ml for 3 days and were pulsed with 0.5 Ci/well [3H]thymidine for 6 h before harvest. H-RS cells dramatically inhibited the prolif- eration of anti-CD3-stimulated T cells. D, paraformaldehyde-fixed H-RS cells or U937 cells were substituted for mitomycin-C-treated cells. There was a dose-dependent inhibitory effect by H-RS cells on T-cell proliferation. E, anti-CD30-treated KM-H2 cells or anti-CD40-treated KM-H2 cells were used. These cells were previously fixed with paraformaldehyde. Anti- CD30-pretreated KM-H2 cells had no effect on anti-CD3-induced T-cell proliferation, whereas the anti-CD40-treated KM-H2 cells preserved its inhibitory effect. C, T cells only; K, KM-H2 cells; H, HDLM cells; L, L428 cells; U, U937 cells; 30-K, anti-CD30-pretreated KM-H2 cells; 40-K, anti-CD40-pretreated KM-H2 cells.

Article Snippet: Immunostaining was performed with goat polyclonal primary antibodies specific for human CD30 (R&D, Minneapolis, MN), followed by incubation with rabbit polyclonal antigoat IgG antibody conjugated to horseradish peroxidase (Santa Cruz Biotechnology, Santa Cruz, CA).

Techniques: Inhibition, Expressing, Activation Assay

Journal: Cancer research

Article Title: CD30 is involved in inhibition of T-cell proliferation by Hodgkin's Reed-Sternberg cells.

doi: 10.1158/0008-5472.can-03-1337

Figure Lengend Snippet: Fig. 3. CD30-Fc fusion protein inhibits anti-CD3-stimulated T-cell proliferation and addition of soluble CD153 restores proliferation. A, a dose-dependent inhibitory effect of the CD30-Fc fusion protein on T-cell proliferation was noted. B, hCD153-mCD8 or hCD154-mCD8 chimeric proteins (1 or 10 g/ml) were incubated on plates coated with CD30-Fc chimeric protein at 10 g/ml. Soluble hCD153 (10 g/ml), but not hCD154 chimeric protein, was able to rescue T-cell proliferation inhibited by CD30-Fc fusion protein. , CD30-Fc; , IgG.

Article Snippet: Immunostaining was performed with goat polyclonal primary antibodies specific for human CD30 (R&D, Minneapolis, MN), followed by incubation with rabbit polyclonal antigoat IgG antibody conjugated to horseradish peroxidase (Santa Cruz Biotechnology, Santa Cruz, CA).

Techniques: Incubation

Journal: Cancer research

Article Title: CD30 is involved in inhibition of T-cell proliferation by Hodgkin's Reed-Sternberg cells.

doi: 10.1158/0008-5472.can-03-1337

Figure Lengend Snippet: Fig. 4. IL-2 production by anti-CD3-stimulated T cells is inhibited by H-RS cells or CD30-Fc fusion protein. A, a standard coculture was performed as described in “Materials and Methods”; the supernatant was collected, and levels of IL-2 were measured. KM-H2 cells inhibited IL-2 production by anti-CD3-treated T cells. However, the inhibition was absent if anti-CD30-treated KM-H2 cells were used. B, addition of IL-2 (1 ng/ml) reversed the inhibition of T-cell proliferation by KM-H2 cells. Numbers denote K/T ratios. C, different concentrations of CD30-Fc fusion protein were substituted for KM-H2 cells. A dose effect on the inhibition of IL-2 production was observed. D, addition of IL-2 (1 ng/ml) effectively rescued the inhibited T-cell proliferation by CD30-Fc. Note the dose- dependent inhibition of T-cell proliferation by CD30-Fc protein. T, T cells; K, KM-H2 cells; 30-K, anti-CD30 pretreated KM-H2 cells. For B and D, black bar represents with IL-2 and gray bar represents without IL-2.

Article Snippet: Immunostaining was performed with goat polyclonal primary antibodies specific for human CD30 (R&D, Minneapolis, MN), followed by incubation with rabbit polyclonal antigoat IgG antibody conjugated to horseradish peroxidase (Santa Cruz Biotechnology, Santa Cruz, CA).

Techniques: Inhibition

Journal: Cancer research

Article Title: CD30 is involved in inhibition of T-cell proliferation by Hodgkin's Reed-Sternberg cells.

doi: 10.1158/0008-5472.can-03-1337

Figure Lengend Snippet: Fig. 5. CD25 and CD26 expression in anti-CD3-stimulated T cells is inhibited by KM-H2 cells or CD30-Fc fusion protein. A, expression of CD25 and CD26 on T cells was analyzed by fluorescence-activated cell sorting. All histograms were gated on CD3 cells. The gray areas denote the intensity of staining of T cells with control antibodies. The white areas denote T cells stained with either anti-CD25 or anti-CD26. In I and III, KM-H2 cells were added to coculture. In II and IV, anti-CD30-treated KM-H2 cells were used. The expression of CD25 and CD26 by anti-CD3-stimulated T cells was similar to those in II and IV, respectively (not shown). B, CD30-Fc fusion protein (middle panel) or human IgG (as control; left panel) was used for study of the effect on CD25 (top panels) and CD26 (bottom panels) expression by anti-CD3-stimulated T cells. Note the down- regulation of CD25 and CD26 by KM-H2 cells and CD30-Fc fusion protein. Addition of IL-2 effectively restored the expression of CD25 by T cells (right panel). 2151

Article Snippet: Immunostaining was performed with goat polyclonal primary antibodies specific for human CD30 (R&D, Minneapolis, MN), followed by incubation with rabbit polyclonal antigoat IgG antibody conjugated to horseradish peroxidase (Santa Cruz Biotechnology, Santa Cruz, CA).

Techniques: Expressing, Fluorescence, FACS, Staining, Control

Journal: Leukemia

Article Title: Notch is an essential upstream regulator of NF-κB and is relevant for survival of Hodgkin and Reed-Sternberg cells.

doi: 10.1038/leu.2011.265

Figure Lengend Snippet: Figure 5 Stimulation of NF-kB signaling in GSI XII treated L540cy cells. (a) HRS cells were seeded in fresh conditioned medium taken from untreated cells after 96 h culture before GSI XII treatment. Viability of cells was determined 24 h later. Immunoblotting of p52 (upper panel), and viability (lower panel) of GSI XII-treated L540cy cells cultured in fresh and conditioned medium. (b) Immunoblotting of p52 and cleaved PARP (left panel) and viability of L540cy cells (right panel) after stimulation through anti-CD30 antibodies (with and without GSI XII treatment).

Article Snippet: Blots Leukemia were incubated with monoclonal rabbit anti-Notch1, anti-p50/ p105, anti-p65 antibodies (Epitomics, Burlingame, CA, USA), anti-cleaved Notch1, rabbit polyclonal anti-p52/p100, antiphospho-p100, anti-RelB, anti-c-Rel, anti-poly-(ADP-ribose) polymerase, anti-cleaved poly-(ADP-ribose) polymerase (Asp214) antibodies (Cell Signaling Technologies, Frankfurt, Germany), anti-human CD30 (R&D Systems) or mouse monoclonal anti-tubulin antibodies (Sigma, Deisenhofen, Germany).

Techniques: Western Blot, Cell Culture