Journal: The Journal of clinical endocrinology and metabolism

Article Title: Association of postpartum thyroid dysfunction with antepartum hormonal and immunological changes.

doi: 10.1210/jc.2002-021219

Figure Lengend Snippet: FIG. 1. Graphical output from the FACScan analyzer (Becton Dickinson and Co.). Before analysis lymphocytes were stained with R-phyco- erythrin-Cy5 mouse antihuman CD3 fluorescein isothiocyanate-conjugated mouse antihuman CD4 together with either phycoerythrin- conjugated rat antihuman IL-4 or phycoerythrin-conjugated mouse antihuman IFN-. The output was gated on the basis of side scatter and CD3 staining. Shown are control cells in the left panels and stimulated cells in the right panels. Lymphocytes gated for right-angle light scatter and CD3 staining (a and b), CD4 expression (c and d), IL-4 expression (c–f), and IFN- expression (e and f) are shown.

Article Snippet: The culture plate was placed in a humidified incubator at 37 C, gas phase 5% CO2 in air, for 4 h. Aliquots (350 l) from the stimulated and unstimulated wells were transferred into fluorescenceactivated cell sorter (FACS) tubes and the surface antigen (CD3 and CD4) was stained by adding 5 l conjugated antibody (R-phycoerythrin-Cy5 conjugated mouse antihuman CD3, DAKO Corp. Ltd., Ely, UK; fluorescein isothiocyanate-conjugated mouse antihuman CD4, Serotec Ltd., Oxford, UK) and incubated in the dark for 30 min.

Techniques: Staining, Control, Expressing

Journal: Scientific Reports

Article Title: 5T4-specific chimeric antigen receptor modification promotes the immune efficacy of cytokine-induced killer cells against nasopharyngeal carcinoma stem cell-like cells

doi: 10.1038/s41598-017-04756-9

Figure Lengend Snippet: Generation and identification of CAR gene-modified CIK cells. ( a ) Schematic representation of the anti-h5T4 CAR construct and the control CAR construct. The 5T4-28Z CAR contained, in sequence from the N-terminus to the C-terminus, a CD8α signal peptide (SP), an anti-h5T4 scFv comprising the variable regions of the heavy chain (VH) and light chain (Vκ) fused with a linker, a myc-tag sequence, a CD8α hinge, the CD28 transmembrane (TM) and intracellular (IC) regions followed by the CD3ζ signalling domain. The control CAR comprised a sequence identical to 5T4-28Z except without the scFv region. ZsGreen was used as a reporter gene. ( b ) CIK cells were generated as mentioned in the Methods section. They were transduced with CAR-encoded lentivirus and sorted using FACS according to the GFP expression. The surface expression of CARs on CIK cells was verified using FACS with an Alexa Fluor ® 647-conjugated myc-tag antibody. As controls, matched isotype antibody staining was incorporated. Flow plots are representative of quintuplicate cultures. ( c ) Brightfield and fluorescence images of sorted 5T4-28Z CAR- or C-28Z CAR-expressing CIK cells were captured. Representative images and merged images are displayed.

Article Snippet: The following antibodies were used: anti-hROR1 polyclonal antibody (R&D), anti-hROR1-APC antibody (R&D), anti-h5T4-APC antibody (R&D), anti-h5T4 mAb (clone EPR5529, Abcam), anti-E-cadherin mAb (clone EP700Y, Abcam), anti-human N-cadherin mAb (clone EPR1791-4, Abcam), anti-human Nanog mAb (clone EPR2027(2), Abcam), anti-human Oct4 mAb (clone EPR2054, Abcam), anti-human vimentin mAb (clone EPR3776, Abcam), anti-human sox2 mAb (clone EPR3131, Abcam), anti-human β-catenin mAb (clone E247, Abcam), anti-human Carbonic Anhydrase IX polyclonal antibody (GeneTex), anti-human GAPDH antibody (Proteintech), HRP-conjugated secondary antibodies (Proteintech), diverse fluorochrome-conjugated anti-human CD3, CD8, CD56, NKG2D, CD57, PD-1, CD107α and isotype control antibodies (Pharmingen, BD).

Techniques: Modification, Construct, Control, Sequencing, Generated, Transduction, Expressing, Staining, Fluorescence

Journal: Scientific Reports

Article Title: 5T4-specific chimeric antigen receptor modification promotes the immune efficacy of cytokine-induced killer cells against nasopharyngeal carcinoma stem cell-like cells

doi: 10.1038/s41598-017-04756-9

Figure Lengend Snippet: CIK cells maintain their original phenotypes after transduction of CAR genes. The expression of CD3, CD8, CD56, NKG2D, CD57, and PD-1 on CAR-engineered CIK cells and NT-CIK cells was monitored weekly using FACS with diverse fluorochrome-conjugated antibodies or matched isotype control antibodies. ( a ) Characteristic flow plots showing the expression of phenotypic markers in 5T4-28Z-CIK cells on day 7, 14, and 21 of cultivation are presented. ( b ) Histograms show the mean values ± SD of three independent experiments.

Article Snippet: The following antibodies were used: anti-hROR1 polyclonal antibody (R&D), anti-hROR1-APC antibody (R&D), anti-h5T4-APC antibody (R&D), anti-h5T4 mAb (clone EPR5529, Abcam), anti-E-cadherin mAb (clone EP700Y, Abcam), anti-human N-cadherin mAb (clone EPR1791-4, Abcam), anti-human Nanog mAb (clone EPR2027(2), Abcam), anti-human Oct4 mAb (clone EPR2054, Abcam), anti-human vimentin mAb (clone EPR3776, Abcam), anti-human sox2 mAb (clone EPR3131, Abcam), anti-human β-catenin mAb (clone E247, Abcam), anti-human Carbonic Anhydrase IX polyclonal antibody (GeneTex), anti-human GAPDH antibody (Proteintech), HRP-conjugated secondary antibodies (Proteintech), diverse fluorochrome-conjugated anti-human CD3, CD8, CD56, NKG2D, CD57, PD-1, CD107α and isotype control antibodies (Pharmingen, BD).

Techniques: Transduction, Expressing, Control

Journal: Frontiers in Immunology

Article Title: Long-term adaptive response in COVID-19 vaccine recipients and the effect of a booster dose

doi: 10.3389/fimmu.2023.1123158

Figure Lengend Snippet: Analysis of B cell frequency in infection-naïve and previously infected vaccine recipients over time. (A) Representative flow cytometry pseudocolor plots of spike-specific B cells in pre-pandemic healthy donor (HD), in an infection-naïve and a previously infected subject. (B, C) Percentages of tetramer + on CD19 + B cells (B) and the percentages of memory CD27 + , IgG + CD27 + and IgA + CD27 + B cells on tetramer + B cells (C) are shown for the spike and RBD protein of the Wuhan Hu-1 SARS-CoV-2 and in response to the B.1.1.529.1 (Omicron) spike protein in infection-naïve and previously infected subjects at 9 months after primary vaccination (T3) and 3 months after the booster dose (T4), as well as for pre-pandemic HD. * p-value <0.05 vs infection-naïve and previously infected subjects at T3 and T4; # p-value <0.05 vs infection-naïve subjects at T3. The sample size ( n ) for all B cell analyses is indicated in brackets in panel (B) .

Article Snippet: Live singlets were gated based on 7AAD fluorescence and specific B cells detected using the double discrimination method gated on CD3 - CD14 - 7AAD - and CD19 + cells (CD3 PerCP-Vio700, #130-113-132; CD14 PerCP-Vio700, #130-113-151, Miltenyi).

Techniques: Infection, Flow Cytometry

Journal: Frontiers in Immunology

Article Title: Unveiling spatial complexity in solid tumor immune microenvironments through multiplexed imaging

doi: 10.3389/fimmu.2024.1383932

Figure Lengend Snippet: Immunophenotyping panel for multiplexed tissue imaging of cancer.

Article Snippet: CD3 , REA1151 , 50 , 130-120-267 , FITC (APC) , Miltenyi Biotec.

Techniques: Imaging