Journal: Scientific Reports

Article Title: Frequency and reactivity of antigen-specific T cells were concurrently measured through the combination of artificial antigen-presenting cell, MACS and ELISPOT

doi: 10.1038/s41598-017-16549-1

Figure Lengend Snippet: Reactivity detection of OVA 257–264 -specific CD8 + T cells by the AAPC-microplate method and traditional ELISPOT assay. After AAPC-bead sorting, the cells retained in each well were further incubated with the AAPC-beads for reactivity evaluation. ( a ) IFN-γ secretion in the OVA 257–264 -specific CD8 + T cell population detected by modified ELISPOT after 24-hr stimulation with the two-signal AAPC-beads in the AAPC-microplate. ( b ) IFN-γ secretion in the spleen cells detected by traditional ELISPOT assay after 24-hr stimulation with OVA 257–264 peptide in a 96-well microplate.

Article Snippet: Mouse interferon-γ (IFN-γ) ELISPOT kit (CT317-T2) and human IFN-γ ELISPOT kit (CT230-T2) were from U-cytech Biosciences (Utrecht, Netherlands).

Techniques: Enzyme-linked Immunospot, Incubation, Modification

Journal: Scientific Reports

Article Title: Frequency and reactivity of antigen-specific T cells were concurrently measured through the combination of artificial antigen-presenting cell, MACS and ELISPOT

doi: 10.1038/s41598-017-16549-1

Figure Lengend Snippet: Enumeration and reactivity detection of HBc 18–27 /HBs 183–191 -specific CD8 + T cells by AAPC-microplate. PBMCs from all subjects were detected by the AAPC-microplate method and traditional HLA-A2/HBc 18–27 /HBs 183–191 dimers staining plus flow cytometry. ( a ) Magnetic separation of HBc 18–27 /HBs 183–191 -specific CD8 + T cells in the AAPC-microplate. Magnification is 400 × for each image. ( b ) The correlation coefficient between the AAPC-microplate method and flow cytometry as analyzed by two-tailed Pearson correlation. ( c ) The frequencies of HBc 18–27 /HBs 183–191 -specific CD8 + T cells in the PBMCs from all subjects as detected by the AAPC-microplate method and flow cytometry (FACS). ( d ) The reactivity of HBc 18–27 /HBs 183–191 -specific CD8 + T cells (AST) and whole T cells from the HLA-A2-positve patients with chronic Hepatitis B. HBc 18–27 /HBs 183–191 -specific CD8 + T cells retained in micro well after AAPC-bead sorting were further co-incubated with PHA for 24 hrs in the AAPC-microplate. Meanwhile, the PBMC samples from the patients were also co-incubated with PHA for 24 hrs without AAPC-bead sorting. IFN-γ local detection was performed by ELISPOT assay as described and the percentages of IFN-γ-secreting cells in the sorted AST cells and CD3 + T cell populations were calculated. Data are presented as mean ± SD.

Article Snippet: Mouse interferon-γ (IFN-γ) ELISPOT kit (CT317-T2) and human IFN-γ ELISPOT kit (CT230-T2) were from U-cytech Biosciences (Utrecht, Netherlands).

Techniques: Staining, Flow Cytometry, Two Tailed Test, Incubation, Enzyme-linked Immunospot

Journal: Scientific Reports

Article Title: Frequency and reactivity of antigen-specific T cells were concurrently measured through the combination of artificial antigen-presenting cell, MACS and ELISPOT

doi: 10.1038/s41598-017-16549-1

Figure Lengend Snippet: IFN-γ-positive spots in the AST cell population and whole T cell population from the HLA-A2-positive patient with hepatitis B. ( a ) The IFN-γ-positive spots in the AST cells from ten HLA-A2-positive patients as detected by the AAPC-microplate with PHA stimulation. ( b ) The IFN-γ-positive spots in the T cells from five HLA-A2-positive patients as detected by the traditional ELISPOT assay with PHA stimulation.

Article Snippet: Mouse interferon-γ (IFN-γ) ELISPOT kit (CT317-T2) and human IFN-γ ELISPOT kit (CT230-T2) were from U-cytech Biosciences (Utrecht, Netherlands).

Techniques: Enzyme-linked Immunospot

Journal: Cellular immunology

Article Title: In vitro ovarian tumor-conditioned CD163+ human macrophages retain phagocytic response to CD47 blockade.

doi: 10.1016/j.cellimm.2025.104932

Figure Lengend Snippet: Fig. 2. Tumor conditioning induces macrophage CD163 and IL-10 gene expression. Tumor-conditioned human monocyte-derived macrophages (M(A2780)) were generated and their gene expression was compared to control macrophages by RT-qPCR. A-F: Presents gene expression of polarization markers (CD163, IL-10, and TNF-α) and phagocytosis checkpoints (SIRPα, LILRB1, and Siglec-10) in tumor-associated macrophages compared to controls. Statistics: comparisons were made against controls using paired t-tests and a significance level of p = 0.05. When indicated by a logarithmic y-axis, comparisons were made on log-transformed data. Data are presented as individual data points with mean ± SD. N = 4.

Article Snippet: The plates were read at 450/ 620 nm and a standard curve ranging from 0.625 to 8 μg/L was prepared using recombinant human LILRB1 (R&D systems, 8989-T2). sCD206 (samples diluted 1:5) [34], and sSIRPα (samples diluted 1:10) [35] were measured as previously reported in detail.

Techniques: Gene Expression, Derivative Assay, Generated, Control, Quantitative RT-PCR, Transformation Assay

Journal: Cellular immunology

Article Title: In vitro ovarian tumor-conditioned CD163+ human macrophages retain phagocytic response to CD47 blockade.

doi: 10.1016/j.cellimm.2025.104932

Figure Lengend Snippet: Fig. 3. Tumor cell conditioning induces CD163, CD206, CD80 and LILRB1 on the membrane protein level. Tumor-conditioned human monocyte-derived macrophages (M(A2780)) were generated and their membrane protein expression compared to control macrophages by flow cytometry. The gating strategy is presented in Supplementary fig. 1. A-G: Presents median fluorescence intensities (MFI) of polarization markers (CD163, CD206, and CD80) and MFI values and percent macrophages positive for phagocytosis checkpoints (LILRB1, and Siglec-10). Statistics: comparisons were made against controls using paired t-tests and a significance level of p = 0.05. When indicated by a logarithmic y-axis, comparisons were made on log-transformed data. Data are presented as individual data points with mean ± SD. N = 9.

Article Snippet: The plates were read at 450/ 620 nm and a standard curve ranging from 0.625 to 8 μg/L was prepared using recombinant human LILRB1 (R&D systems, 8989-T2). sCD206 (samples diluted 1:5) [34], and sSIRPα (samples diluted 1:10) [35] were measured as previously reported in detail.

Techniques: Membrane, Derivative Assay, Generated, Expressing, Control, Flow Cytometry, Fluorescence, Transformation Assay

Journal: Nucleic Acids Research

Article Title: Polymerization retardation isothermal amplification (PRIA): a strategy enables sensitively quantify genome-wide 5-methylcytosine oxides rapidly on handy instruments with nanoscale sample input

doi: 10.1093/nar/gkz704

Figure Lengend Snippet: Principle of PRIA assay for genome-wide 5hmC, 5fC and 5caC detection. ( A ) Universal adaptor T 2 ligated genomic ssDNA fragments with or without labeling. ( B ) Chemical labeling of 5fC-DNA, 5hmC-DNA and 5caC-DNA by PHPA, T4- β GT/UDP-glucose and 3-CPBA, and LY, respectively. ( C ) Isothermal replication-scission amplification reaction of the ssDNA fragments on a laboratory rt-qPCR instrument. ( D ) Data analysis for screening credible slope.

Article Snippet: Klenow fragment polymerase (KF, 3′ → 5′ exo-), Nt.BsmAI NEase, T4 Phage β -glucosyltransferase (T4- β GT), UDP-glucose, Cutsmart buffer (20 mM Tris-acetate, 50 mM potassium acetate, 10 mM magnesium acetate and 0.1 mg ml –1 BSA, pH 7.9), NEB buffer 2 (10 mM Tris–HCl, 10 mM MgCl 2 , 50 mM NaCl and 1 mM dithiothreitol, pH 7.9), NEB buffer 4 (50 mM potassium acetate, 20 mM Tris-acetate, 10 mM magnesium acetate and 1 mM dithiothreitol, pH 7.9), the NEBNext Ultra End Repair/dA-Tailing Module, the NEBNext Ultra Ligation Module and NEBNext Singleplex Oligos for Illumina were purchased from New England Biolabs (USA).

Techniques: Genome Wide, Labeling, Amplification, Quantitative RT-PCR

Journal: Nucleic Acids Research

Article Title: Polymerization retardation isothermal amplification (PRIA): a strategy enables sensitively quantify genome-wide 5-methylcytosine oxides rapidly on handy instruments with nanoscale sample input

doi: 10.1093/nar/gkz704

Figure Lengend Snippet: Feasibility of PRIA for the detection of 5hmC. ( A ) MALDI-TOF MS characterization of 5hmC-DNA after labeling with UDP-glucose and 3-CPBA. ( B ) Time-dependent fluorescence spectra of the isothermal replication-scission amplification reactions with 5hmC-DNA before and after labeling with UDP-glucose and 3-CPBA. ( C ) Histogram of Δ S for the isothermal replication-scission amplification reactions with C-DNA, 5mC-DNA, 5hmC-DNA, 5fC-DNA and 5caC-DNA before and after their reactions with T4- β GT/UDP-glucose and 3-CPBA. ( D ) Histogram of Δ S for the isothermal replication-scission amplification reactions with diverse proportions of CPBA-5gmC-DNA, and plot of Δ S versus CPBA-5gmC-DNA content.

Article Snippet: Klenow fragment polymerase (KF, 3′ → 5′ exo-), Nt.BsmAI NEase, T4 Phage β -glucosyltransferase (T4- β GT), UDP-glucose, Cutsmart buffer (20 mM Tris-acetate, 50 mM potassium acetate, 10 mM magnesium acetate and 0.1 mg ml –1 BSA, pH 7.9), NEB buffer 2 (10 mM Tris–HCl, 10 mM MgCl 2 , 50 mM NaCl and 1 mM dithiothreitol, pH 7.9), NEB buffer 4 (50 mM potassium acetate, 20 mM Tris-acetate, 10 mM magnesium acetate and 1 mM dithiothreitol, pH 7.9), the NEBNext Ultra End Repair/dA-Tailing Module, the NEBNext Ultra Ligation Module and NEBNext Singleplex Oligos for Illumina were purchased from New England Biolabs (USA).

Techniques: Labeling, Fluorescence, Amplification