Journal: Human Gene Therapy

Article Title: A Short Polypeptide from the Herpes Simplex Virus Type 2 ICP10 Gene Can Induce Antigen Aggregation and Autophagosomal Degradation for Enhanced Immune Presentation

doi: 10.1089/hum.2010.080

Figure Lengend Snippet: ELISPOT assays of immune responses to class I and II OVA peptides. Mice were vaccinated with the indicated constructs twice before spleens were taken for ELISPOT assays as described in Materials and Methods. (A) Quantification of IFN-γ ELISPOT from purified CD4+ T cells after stimulation with class II OVA peptide. (B) Results of quantification of IFN-γ ELISPOT from purified splenocytes after stimulation with class I OVA peptide. Star, p < 0.01 as compared with vaccinations of B16 or B16-pOVA; cross, p < 0.05 as compared with vaccination of B16.

Article Snippet: To separate CD4 + cells, Ficoll-purified spleen cells were passed through murine CD4 + T cell enrichment columns (BD Biosciences, San Jose, CA).

Techniques: Enzyme-linked Immunospot, Construct, Purification

Journal:

Article Title: Activation of antigen-presenting cells by microbial products breaks self tolerance and induces autoimmune disease

doi: 10.1172/JCI200419388

Figure Lengend Snippet: Flow cytometry analysis of thymocytes and peripheral T cells in 5B6 transgenic B10.S mice. (A and B) T cells from thymi (A) and spleens (B) of mice were stained with the indicated antibodies (PE- or allophycocyanin-conjugated anti-CD4, FITC-conjugated anti-CD8, and PE-conjugated anti–TCR Vβ6). Dot plots representing two-color flow cytometry analysis of 5B6 transgenic mice with or without RAG-2 deficiency (TG/RAG-2–/– or TG, respectively) and nontransgenic littermates (NLM) are shown. Numbers in quadrants refer to percentages of gated cell populations. Total numbers of spleen cells are shown above dot plots.

Article Snippet: Spleen cell samples from 5B6 transgenic mice were enriched for CD4 + T cells by negative selection using CD4 T cell enrichment columns (Cytovax, Edmonton, Alberta, Canada) according to the manufacturer’s protocol.

Techniques: Flow Cytometry, Transgenic Assay, Staining

Journal:

Article Title: Activation of antigen-presenting cells by microbial products breaks self tolerance and induces autoimmune disease

doi: 10.1172/JCI200419388

Figure Lengend Snippet: Responses of 5B6 transgenic T cells to PLP139–151. (A) Proliferative response of 5B6 transgenic T cells to PLP139–151 presented by DAS cells. CD4-enriched T cell samples from 5B6 transgenic SJL and B10.S mice were stimulated with the indicated numbers of PLP139–151–pulsed DAS cells. T cell proliferation was assessed by [3H]thymidine incorporation assay. The mean cpm ± SD of triplicate cultures are shown. (B) Proliferative response of T cells from 5B6 transgenic B10.S or SJL mice to PLP139–151 presented by syngeneic APCs. Splenocyte samples from unimmunized 5B6 transgenic and nontransgenic littermate (NLM) B10.S or SJL mice were enriched for CD4+ T cells and were cultured with irradiated splenocytes from nontransgenic littermates in the presence of the indicated concentrations of PLP139–151 or control peptide PLP178–191. Proliferative responses were determined by [3H]thymidine incorporation assay. The mean cpm ± SD of triplicate cultures of one experiment representative of three are shown. (C) IL-2 response to PLP139–151 of T cells from 5B6 transgenic B10.S or SJL mice. Supernatants from cultures in B were assayed in duplicate by ELISA for cytokine production. Representative data from one of three experiments are shown. (D) INF-γ response of spleen cells from 5B6 transgenic B10.S or SJL mice to PLP139–151 stimulation. Culture supernatants from 5B6 transgenic SJL or B10.S whole spleen cells stimulated with the indicated concentrations of PLP139–151 were assayed in duplicate by ELISA for INF-γ production.

Article Snippet: Spleen cell samples from 5B6 transgenic mice were enriched for CD4 + T cells by negative selection using CD4 T cell enrichment columns (Cytovax, Edmonton, Alberta, Canada) according to the manufacturer’s protocol.

Techniques: Transgenic Assay, Thymidine Incorporation Assay, Cell Culture, Irradiation, Enzyme-linked Immunosorbent Assay