Journal: Croatian Medical Journal

Article Title: Ornidazole suppresses CD133 + melanoma stem cells via inhibiting hedgehog signaling pathway and inducing multiple death pathways in a mouse model

doi: 10.3325/cmj.2022.63.461

Figure Lengend Snippet: Results of Crystal Violet and MTT assays. (A) Cellular viability was assessed with Crystal Violet staining. Melanoma cancer cells were treated with the denoted concentrations of ornidazole for 48 h. (B-D) Ornidazole inhibits B16F10 melanoma cancer cells viability as determined by MTT assay. Melanoma cancer cells were treated with different concentrations of ornidazole for (B) 24 h, (C) 48 h, (D) 72 h. Results were presented as mean ± standard deviation of three independent experiments. Each group with different concentration of ornidazole was compared with the control group (one-way ANOVA with Dunnet's test for multiple comparisons, * P < 0.05; ** P < 0.01; *** P < 0.001).

Article Snippet: B16F10 mouse melanoma cancer cell line was purchased from ATCC (CRL-6475, Manassas, VA, USA).

Techniques: Staining, MTT Assay, Standard Deviation, Concentration Assay, Control

Journal: Croatian Medical Journal

Article Title: Ornidazole suppresses CD133 + melanoma stem cells via inhibiting hedgehog signaling pathway and inducing multiple death pathways in a mouse model

doi: 10.3325/cmj.2022.63.461

Figure Lengend Snippet: Wound-healing and Comet assays. The results are expressed as percentage of wound closure, and the area measured at time zero was considered 0% (A) Ornidazole inhibits B16F10 melanoma cancer cells migration as determined by wound-healing analysis. (B) Wound closure rates (%) at 8 h and 24 h. Ornidazole (400 μg/mL, 800 μg/mL, 1200 μg/mL) inhibited cell migration by 15%, 60%, and 96%, respectively, in B16F10 cells after 24 hours (one-way ANOVA followed by Bonferroni testing for multiple comparisons. Data are shown as the mean ± standard deviation, P < 0.05) (C) DNA damage as measured by the Comet assay as a result of treatment with different concentrations of ornidazole (400 μg/mL, 800 μg/mL, 1200 μg/mL) for 24, 48, and 72 hours. Olive moment = (tail mean-head mean) x % of DNA in the tail (D) Mean tail moment (μm) represents the damage distribution in the attached cells. The experiment was done in triplicates, and data are expressed as mean ± standard deviation. Each group was compared with the control group (two-way ANOVA with multiple comparisons, Bonferroni test was used for post hoc analysis, *** P < 0.001).

Article Snippet: B16F10 mouse melanoma cancer cell line was purchased from ATCC (CRL-6475, Manassas, VA, USA).

Techniques: Migration, Standard Deviation, Single Cell Gel Electrophoresis, Control

Journal: Croatian Medical Journal

Article Title: Ornidazole suppresses CD133 + melanoma stem cells via inhibiting hedgehog signaling pathway and inducing multiple death pathways in a mouse model

doi: 10.3325/cmj.2022.63.461

Figure Lengend Snippet: (A) Mice that were injected unsorted B16F10 cells and tumors resected from ornidazole-treated and control mice that received unsorted cell injection. (B) Mice that were injected CD133 + B16F10 cells. (C) Mice that were injected CD133 - B16F10 cells and tumors resected from ornidazole-treated and control mice that received CD133 - cell injection. (D) After ornidazole treatment, average tumor volume measurements of the unsorted group (*** P < 0.001), (E) CD133 + (4th day ** P < 0.01, 9th-12th day *** P < 0.001), and (F) CD133 - groups (2nd day * P < 0.5, 9th-12th day *** P < 0.001). All groups were compared with the control group (two-way ANOVA with Bonferroni correction for multiple comparisons).

Article Snippet: B16F10 mouse melanoma cancer cell line was purchased from ATCC (CRL-6475, Manassas, VA, USA).

Techniques: Injection, Control

Journal: Croatian Medical Journal

Article Title: Ornidazole suppresses CD133 + melanoma stem cells via inhibiting hedgehog signaling pathway and inducing multiple death pathways in a mouse model

doi: 10.3325/cmj.2022.63.461

Figure Lengend Snippet: Gene and relative protein expression levels of hedgehog signaling pathway. (A) Effect of ornidazole on hedgehog signaling pathway in unsorted, CD133 + , and CD133 - cells was assessed with real time-polymerase chain reaction. Tumor cells were isolated from mice. Each group was compared with the control group. Data are shown as mean ± standard deviation from three independent experiments (one-way ANOVA with Bonferroni corrections for multiple comparisons, *** P < 0.001, ** P < 0.01 (n = 6/group). (B) The fold-change levels of protein expression in ELISA assay of the B16F10 cells treated with ornidazole. The experiments were performed in duplicate. Each group was compared with the control group. Results are presented as mean ± standard deviation of three independent experiments (two-way ANOVA with Bonferroni testing for multiple comparisons, *** P < 0.001, ** P < 0.01, n = 6 per group). Shh – Sonic hedgehog, PTCH1 – patched-1, SMO – smoothened, GLI1 – glioma-associated oncogene homologue-1, CD133 – also known as AC133 and prominin-1.

Article Snippet: B16F10 mouse melanoma cancer cell line was purchased from ATCC (CRL-6475, Manassas, VA, USA).

Techniques: Expressing, Real-time Polymerase Chain Reaction, Isolation, Control, Standard Deviation, Enzyme-linked Immunosorbent Assay

Journal:

Article Title: Aggravation of Anti-Myeloperoxidase Antibody-Induced Glomerulonephritis by Bacterial Lipopolysaccharide

doi:

Figure Lengend Snippet: Immunophenotyping of Intraglomerular Inflammatory Cell Infiltrate at 6 Days after Anti-TNF-α Antibody or Isotype Control Antibody (L2-3D9) Treatment

Article Snippet: To investigate the effects of TNF-α depletion on disease induction, groups of mice received a single intraperitoneal dose of the anti-murine TNF-α antibody TN3 (500 μg/mouse, endotoxin concentration <10 pg/ml), a complementarity-determining regions-grafted murine IgG2a ( n = 8; kindly provided by Celltech, Slough, UK) or isotype control (mAb L2-3D9, endotoxin concentration <10 pg/ml, n = 7), 19,20 in sterile PBS, 24 hours before anti-MPO IgG and LPS (0.5 μg/g) administration.

Techniques: Control

Journal: The Journal of Clinical Investigation

Article Title: POLQ inhibition elicits an immune response in homologous recombination–deficient pancreatic adenocarcinoma via cGAS/STING signaling

doi: 10.1172/JCI165934

Figure Lengend Snippet: ( A ) Representative images of KPC and KPC- Brca2 –/– shCtrl and shP OLQ tumor sections stained for CD4 + , CD8 + T cells, and F4/80 + cells by IHC. Scale bar: 50 μm. ( B ) Quantification of CD4 + (upper), CD8 + T cells (middle), and F4/80 + cells (lower) from A . Each point on the graph represents 1 mouse ( n = 5 mice/group). HPF, high-power field. ( C ) CD4 + cells as a percentage of CD3 + T cells during flow cytometry of KPC- Brca2 –/– tumors, shCtrl, and shP OLQ ( n = 10 tumors). ( D ) CD8 + cells as a percentage of CD3 + T cells during flow cytometry of KPC- Brca2 –/– tumors with shCtrl and shP OLQ ( n = 10 tumors). ( E ) Representative images of coIF staining of CD8 (red), Granzyme B (GrB; green), and DAPI (blue) in KPC- Brca2 –/– shCtrl and shP OLQ tumors. Scale bar: 20 μm. ( F and G ) Quantification of the percent of CD8 + cells ( F ) or percent of CD8 + and GrB + double-positive cells ( G ) from E ( n = 5 tumors). ( H ) CD11b + cells as a percentage of CD45 + cells during flow cytometry of KPC- Brca2 –/– tumors with shCtrl and shPOLQ ( n = 10 tumors). ( I ) CD206 – MHC II + CD11b + cells as a percentage of CD45 + cells during flow cytometry of KPC- Brca2 –/– tumors with shCtrl, and shP OLQ ( n = 10 tumors). ( J ) CD206 + MHC II – CD11b + cells as a percentage of CD45 + cells during flow cytometry of KPC- Brca2 –/– tumors with shCtrl and shP OLQ ( n = 10 tumors). Data are representative of at least 3 independent experiments. * P < 0.05, ** P < 0.01, *** P < 0.005. Error bars indicate the mean ± SEM.

Article Snippet: Single cells from tumor digestion were incubated with 2 μg/mL anti-mouse CD3 (BD Biosciences, 553057), 1 μg/mL anti-mouse CD28 (BD Biosciences, 553294), and 1 μL/mL Cell Stimulation Cocktail (eBioscience) for 4 hours at 37°C in RPMI with 10% FBS.

Techniques: Staining, Flow Cytometry

Journal: Immunity

Article Title: Aryl Hydrocarbon Receptor Controls Monocyte Differentiation into Dendritic Cells versus Macrophages.

doi: 10.1016/j.immuni.2017.08.016

Figure Lengend Snippet: Figure 2. IRF4 and MAFB Are Essential for mo-DC and mo-Mac Differentiation (A) Volcano plot showing the fold change and significance of transcription factor genes between ascites mo-DCs and mo-Macs. Genes not expressed in in vitro- generated mo-DCs or mo-Macs were filtered out. Adjusted p values determined by differential expression analysis. (B) Cell-sorted ascites mo-DCs and mo-Macs and blood CD14+ monocytes (Mono) were analyzed by immunoblot. Representative of four independent experiments. (C) CD14+ monocytes were cultured with MCSF, IL-4, and TNF-a for 3 hr, 6 hr, 12 hr, or 24 hr or processed directly after isolation (0). IRF4 and MAFB expression were analyzed by RT-qPCR in total cells. Each color represents an individual donor (n = 5 in 3 independent experiments). (D) CD14+ monocytes were cultured with MCSF, IL-4, and TNF-a for 5 days. Total cells were lysed at different days and analyzed by immunoblot. Representative of five independent experiments. (E–H) Monocytes were infected at day 0 with lentivirus containing shRNA against IRF4 (E and F) or MAFB (G and H) or control shRNA. After 5 days of culture, cells were analyzed by immunoblot (E and G) or by flow cytometry (F and H).

Article Snippet: REAGENT or RESOURCE SOURCE IDENTIFIER Biological Samples Ascites from ovarian cancer patients Hospital of Institut Curie (Paris) N/A Ascites from breast cancer patients Hospital of Institut Curie (Paris) N/A Peripheral blood from buffy coats Etablissement français du Sang (Paris) N/A Chemicals, Peptides, and Recombinant Proteins Stemregenin-1 Cayman chemicals SSY261 6-Formylindolo(3,2-b)carbazole Enzo Life Sciences BML-GR206-0100 Critical Commercial Assays PrimeFlow RNA Assay eBioscience Cat: # 88-18005-210 RNAeasy micro kit QIAGEN Cat: #74004 Deposited Data Affymetrix data GEO GSE102046 Single-cell RNA-seq data GEO GSE103544 Experimental Models: Organisms/Strains Mouse: B6.129-Ahrtm1Gonz/Nci Fernandez-Salguero et al., 1995 RRID:IMSR_NCIMR:01XC3 Oligonucleotides Primer IRF4 Thermo Fisher Scientific Hs01056533_m1 Primer CYP1A1 Thermo Fisher Scientific Hs01054797_g1 Primer MAFB Thermo Fisher Scientific Hs00271378_s1 Primer PRDM1 Thermo Fisher Scientific Hs00153357_m1 Software and Algorithms Cell Ranger Single Cell Software Suite (v1.3.1) Zheng et al., 2017 https://support.10xgenomics.com/single-cell- gene-expression/software/pipelines/latest/rkit Seurat package (v1.4.0.14) Satija et al., 2015 https://github.com/satijalab/seurat Ade4 package Chessel et al., 2004 http://pbil.univ-lyon1.fr/ade4/ EMA package Servant et al., 2010 http://bioinfo-out.curie.fr/projects/ema/ ArrayQualityMetrics package Kauffmann et al., 2009 https://bioconductor.org/packages/release/ bioc/html/arrayQualityMetrics.html oligo package Carvalho and Irizarry, 2010 https://www.bioconductor.org/packages/ release/bioc/html/oligo.html limma package Smyth, 2004 https://bioconductor.org/packages/release/ bioc/html/limma.html FlowJo (v.9.9.5) FlowJo LLC https://www.flowjo.com/

Techniques: In Vitro, Generated, Quantitative Proteomics, Western Blot, Cell Culture, Isolation, Expressing, Quantitative RT-PCR, Infection, shRNA, Control, Cytometry

Journal: Contrast Media & Molecular Imaging

Article Title: 99m Tc-Labeled LyP-1 for SPECT Imaging of Triple Negative Breast Cancer

doi: 10.1155/2019/9502712

Figure Lengend Snippet: Cytotoxic effects of LyP-1 and ARAL on 4T1 cells at concentrations from 0.1 to 50 μ M after incubation for 24 h.

Article Snippet: 4T1 murine breast carcinoma cells were purchased from American Type Culture Collection (Manassas, VA, USA) and maintained in RPMI-1640 supplemented with 10% fetal bovine serum (FBS, Gibco, Grand Island, NY, USA).

Techniques: Incubation

Journal: Contrast Media & Molecular Imaging

Article Title: 99m Tc-Labeled LyP-1 for SPECT Imaging of Triple Negative Breast Cancer

doi: 10.1155/2019/9502712

Figure Lengend Snippet: Cellular uptake of LyP-1-FITC and ARAL-FITC by 4T1 cells was examined by flow cytometry. Cells were treated with the drug-free medium (a), ARAL-FITC (b), or LyP-1-FITC (c) at a concentration of 10 μ M for 2 h, and the statistical analysis results are shown in (d). Cells treated with 10 μ M ARAL-FITC (e) and LyP-1-FITC (f) were analyzed after incubation for 3 h, and the difference was compared in (g). The relative fluorescence signals of cells treated with ARAL-FITC (h) or LyP-1-FITC (i) were compared for 2-3 h incubation, respectively. ∗∗ < 0.01.

Article Snippet: 4T1 murine breast carcinoma cells were purchased from American Type Culture Collection (Manassas, VA, USA) and maintained in RPMI-1640 supplemented with 10% fetal bovine serum (FBS, Gibco, Grand Island, NY, USA).

Techniques: Flow Cytometry, Concentration Assay, Incubation, Fluorescence

Journal: Contrast Media & Molecular Imaging

Article Title: 99m Tc-Labeled LyP-1 for SPECT Imaging of Triple Negative Breast Cancer

doi: 10.1155/2019/9502712

Figure Lengend Snippet: Laser confocal microscopy images of 4T1 cells incubated with ARAL-FITC or LyP-1-FITC at a concentration of 10 μ M for 3 h.

Article Snippet: 4T1 murine breast carcinoma cells were purchased from American Type Culture Collection (Manassas, VA, USA) and maintained in RPMI-1640 supplemented with 10% fetal bovine serum (FBS, Gibco, Grand Island, NY, USA).

Techniques: Confocal Microscopy, Incubation, Concentration Assay

Journal: Contrast Media & Molecular Imaging

Article Title: 99m Tc-Labeled LyP-1 for SPECT Imaging of Triple Negative Breast Cancer

doi: 10.1155/2019/9502712

Figure Lengend Snippet: SPECT images of 4T1 cells incubated with 99m Tc-LyP-1 and 99m Tc-ARAL at different radiodoses for 4 h in vitro SPECT images (a) and their relative SPECT signal intensities (b) of 4T1 cells treated with 99m Tc-LyP-1 or 99m Tc-ARAL at 99m Tc radioactive concentrations of 25, 50, 100, 200, and 400 μ Ci/mL, respectively. ∗ < 0.05, ∗∗ < 0.01, and ∗∗∗ < 0.001.

Article Snippet: 4T1 murine breast carcinoma cells were purchased from American Type Culture Collection (Manassas, VA, USA) and maintained in RPMI-1640 supplemented with 10% fetal bovine serum (FBS, Gibco, Grand Island, NY, USA).

Techniques: Single Photon Emission Computed Tomography, Incubation, In Vitro

Journal: Contrast Media & Molecular Imaging

Article Title: 99m Tc-Labeled LyP-1 for SPECT Imaging of Triple Negative Breast Cancer

doi: 10.1155/2019/9502712

Figure Lengend Snippet: LyP-1 recognizes tumor lymphatics in a 4T1 mouse model. Colocalization of LyP-1-FITC (green) in 4T1 tumor tissue with CD31 (a) (red, a blood vessel marker) and LYVE-1 (b) (red, a lymphatic endothelial marker). Scale bar, 100 μ m.

Article Snippet: 4T1 murine breast carcinoma cells were purchased from American Type Culture Collection (Manassas, VA, USA) and maintained in RPMI-1640 supplemented with 10% fetal bovine serum (FBS, Gibco, Grand Island, NY, USA).

Techniques: Marker

Journal: Contrast Media & Molecular Imaging

Article Title: 99m Tc-Labeled LyP-1 for SPECT Imaging of Triple Negative Breast Cancer

doi: 10.1155/2019/9502712

Figure Lengend Snippet: SPECT imaging of 99m Tc-LyP-1 in vivo. SPECT images of nude mice bearing 4T1 xenografted tumors at different time points after intravenous injection of 99m Tc-LyP-1 (a). The tumor-to-muscle (T/M) ratios of 99m Tc-LyP-1 and 99m Tc-ARAL were compared at 0.5 min, 0.5 h, 1 h, 2 h, 4 h, and 6 h (b). The relative SPECT signal intensities of ex vivo tumors and major organs (c) at 6 h postinjection of 99m Tc-LyP-1 or 99m Tc-ARAL. The ratio of tumor/liver (T/L) and tumor/kidney (T/K) of 99m Tc-LyP-1 (d) and 99m Tc-ARAL (e) was compared at 6 h postinjection. ∗∗∗ < 0.001.

Article Snippet: 4T1 murine breast carcinoma cells were purchased from American Type Culture Collection (Manassas, VA, USA) and maintained in RPMI-1640 supplemented with 10% fetal bovine serum (FBS, Gibco, Grand Island, NY, USA).

Techniques: Single Photon Emission Computed Tomography, Imaging, In Vivo, Injection, Ex Vivo

Journal:

Article Title: Transient blockade of CD40 ligand dissociates pathogenic from protective mucosal immunity

doi: 10.1172/JCI13720

Figure Lengend Snippet: CTL priming by systemic or oral OVA requires CD40L signaling. (a) A single intraperitoneal injection of 250 μg of control mAb 6C8 (squares) or anti-CD40L mAb MR1 (circles) was given to B6 mice (n = 4 in each group), 1 day before challenge with 2 × 107 intravenous OVA-coated H-2Kbm-1 splenocytes to prime CTLs. After 14 days, mice were killed and their splenocytes were tested for CTL activity, expressed as OVA-specific lysis for representative individual mice. Primed splenocytes as effectors (E) were tested against 51Cr-loaded cells as targets (T). (b) The same doses of 6C8 or MR1 were given to mice (n = 12 in each group) that were then fed 20 mg OVA on three alternating days. After 14 days, without further priming, mice were killed and their splenocytes were tested for CTL activity, this time expressed as lytic units per spleen for individual mice.

Article Snippet: Two hours prior to the first OVA treatment, mice were injected with a single intraperitoneal dose of 250 μg hamster IgG1 anti-mouse CD40L mAb (MR1; American Type Culture Collection, Rockville, Maryland, USA) or the control hamster IgG1 mAb 6C8, which is specific for human Bcl-2 ( 28 ).

Techniques: Injection, Control, Activity Assay, Lysis

Journal:

Article Title: Transient blockade of CD40 ligand dissociates pathogenic from protective mucosal immunity

doi: 10.1172/JCI13720

Figure Lengend Snippet: Anti-CD40L treatment prevents induction of diabetes by oral OVA in RIP-OVAlo mice. RIP-OVAlo mice bearing OT-I and OT-II cells were injected intraperitoneally with 250 μg control mAb 6C8 (squares) or anti-CD40L mAb MR1 (circles). To mimic low-dose oral tolerance regimens, mice were then fed 0.5 mg OVA on five alternating days. Blood glucose was measured 12 days after the start of feeding; values above 12 mmol/l were considered diagnostic of diabetes. Data on individual mice are pooled from two experiments.

Article Snippet: Two hours prior to the first OVA treatment, mice were injected with a single intraperitoneal dose of 250 μg hamster IgG1 anti-mouse CD40L mAb (MR1; American Type Culture Collection, Rockville, Maryland, USA) or the control hamster IgG1 mAb 6C8, which is specific for human Bcl-2 ( 28 ).

Techniques: Injection, Control, Diagnostic Assay

Journal:

Article Title: Transient blockade of CD40 ligand dissociates pathogenic from protective mucosal immunity

doi: 10.1172/JCI13720

Figure Lengend Snippet: Anti-CD40L treatment does not prevent oral tolerance as measured by suppression of systemic priming of CTLs. CTL activity in response to intravenous priming with OVA-coated splenocytes (a) or to subcutaneous priming with OVA in CFA (b) is similarly suppressed after oral OVA given to mice treated with control mAb 6C8 (squares) and those given anti-CD40L mAb MR1 (circles). Mice were injected with 6C8 or MR1 (250 μg intraperitoneally), then fed either PBS (open symbols) or OVA in PBS (filled symbols). After 14 days, mice were primed systemically. Seven days later they were killed and their splenocytes were recovered for a standard in vitro 51Cr release assay. CTL activity was expressed as percent OVA-specific lysis; the percent of total radioactivity, corrected for background, released from 51Cr-loaded, OVA257-264 peptide-coated target cells (T) by primed effector spleen cells (E). CTL activity plots for individual mice (shown in a and b) were converted into lytic units per spleen from four experiments (c) in which mice received either PBS or 20 mg oral OVA in PBS on three alternating days and were primed as in a, and from two experiments (d) in which mice received either PBS or 0.5 mg oral OVA in PBS on five alternating days and were primed as in b. Horizontal bars are mean values.

Article Snippet: Two hours prior to the first OVA treatment, mice were injected with a single intraperitoneal dose of 250 μg hamster IgG1 anti-mouse CD40L mAb (MR1; American Type Culture Collection, Rockville, Maryland, USA) or the control hamster IgG1 mAb 6C8, which is specific for human Bcl-2 ( 28 ).

Techniques: Activity Assay, Control, Injection, In Vitro, Release Assay, Lysis, Radioactivity

Journal:

Article Title: Transient blockade of CD40 ligand dissociates pathogenic from protective mucosal immunity

doi: 10.1172/JCI13720

Figure Lengend Snippet: Activation and expansion of CTLs by oral OVA requires CD40L. B6 recipient mice congenic for Ly5.1 were adoptively transferred with 3 × 106 transgenic OT-I cells (Ly5.2) and then given control mAb 6C8 or anti-CD40L mAb MR1 (250 μg, intraperitoneally). Mice from each treatment group were then divided into two groups and fed either PBS or 20 mg OVA in PBS on three alternating days. mAb treatment was repeated before the third feeding. Mice were killed 14 days from the start of feeding, and the numbers and phenotype of OT-I cells in their spleens were analyzed by flow cytometry. (a) Dot plots of individual mice show CD44 (left) and CD62L (L-selectin) (right) expression on OT-I cells. The percentage of cells expressing a high level of CD44 or a low level of CD62L is shown in the corresponding quadrant. The number of OT-I cells per spleen (b), CD44 expression as mean fluorescence intensity (MFI) (c), and % CD62Llo OT-I cells (d) in individual recipient mice treated with 6C8 (squares) or MR1 (circles) and then fed PBS (open symbols) or OVA (filled symbols). Data on individual mice are pooled from two experiments.

Article Snippet: Two hours prior to the first OVA treatment, mice were injected with a single intraperitoneal dose of 250 μg hamster IgG1 anti-mouse CD40L mAb (MR1; American Type Culture Collection, Rockville, Maryland, USA) or the control hamster IgG1 mAb 6C8, which is specific for human Bcl-2 ( 28 ).

Techniques: Activation Assay, Transgenic Assay, Control, Flow Cytometry, Expressing, Fluorescence

Journal:

Article Title: Transient blockade of CD40 ligand dissociates pathogenic from protective mucosal immunity

doi: 10.1172/JCI13720

Figure Lengend Snippet: Anti-CD40L treatment does not prevent oral tolerance measured as suppression of systemic priming of (a) T cell proliferation measured as 3H-thymidine (T) incorporation, or (b) IFN-γ production measured as ELIspots. Mice (n = 3 in each group) were injected with control mAb 6C8 or anti-CD40L mAb MR1 (250 μg intraperitoneally), and then fed either PBS or 20 mg OVA in PBS on three alternating days. After 7 days, they were immunized subcutaneously with OVA (0.1 mg) in CFA in the base of tail. Ten days later, spleens and inguinal lymph nodes were harvested for measurement of T cell proliferation (a) and cytokine production (b) in the absence (white bars) or presence (black bars) of 0.1 mg/ml OVA (mean and SD shown for spleen), and sera obtained for measurement of anti-OVA Ab’s (not shown, see text), as described in Methods. 3H-T, 3H-thymidine.

Article Snippet: Two hours prior to the first OVA treatment, mice were injected with a single intraperitoneal dose of 250 μg hamster IgG1 anti-mouse CD40L mAb (MR1; American Type Culture Collection, Rockville, Maryland, USA) or the control hamster IgG1 mAb 6C8, which is specific for human Bcl-2 ( 28 ).

Techniques: Injection, Control