Journal: iScience

Article Title: Wnt antagonism without TGFβ induces rapid MSC chondrogenesis via increasing AJ interactions and restricting lineage commitment

doi: 10.1016/j.isci.2022.105713

Figure Lengend Snippet:

Article Snippet: Human bone marrow MSC , PromoCell , Cat#C-12974.

Techniques: Purification, Blocking Assay, Recombinant, In Situ, Software

Journal: Cell Reports Methods

Article Title: An efficient immunoassay for the B cell help function of SARS-CoV-2-specific memory CD4 + T cells

doi: 10.1016/j.crmeth.2022.100224

Figure Lengend Snippet: BAFF and SCGFβ are majorly implicated in the monocytes-driven enhanced B cell survival (A) Volcano plot shows the concentration difference versus −log10 adjusted p values for the comparison of secreted factors between B + M and B-alone conditions after 9 days. Statistical calculation was done by multiple t test corrected using the Bonferroni-Sidak method. Dotted line represents p = 0.05. Red open circles represent the significantly secreted analyte. (B) Dot plot shows the fold change of various secreted factors in the same conditions as mentioned in (A). Dotted line represents the fold change = 2. Red open circles represent the analyte with fold change of ≥2. Data are representative of n = 10 donors. (C) Flow cytometric plots show the live B cells (CD19 + ) in 6 days B + M cocultures in presence of isotype (mouse IgG) or various blocking conditions. (D) Bar plot shows the count of live B cells in various blocking conditions, mentioned in (C). (E) Percent survival of B cells (relative to B + M) in presence of isotype, αBAFF, or αBAFF + αSCGFβ blocking conditions. Data are shown as mean ± SEM. Data represent the pool of two independent experiments. Statistics are as follows: (D and E) one-way ANOVA followed by Bonferroni’s multiple comparisons test. See also Figure S8 .

Article Snippet: In some B cell survival assay, B cells were supplemented with either the recombinant BAFF (#2149-BF, R&D Systems) or SCGF (#1904-SC, R&D Systems) at various concentrations.

Techniques: Concentration Assay, Comparison, Blocking Assay

Journal: Cell Reports Methods

Article Title: An efficient immunoassay for the B cell help function of SARS-CoV-2-specific memory CD4 + T cells

doi: 10.1016/j.crmeth.2022.100224

Figure Lengend Snippet: Promoting B cell survival by growth factors BAFF and SCGF is not sufficient to replace monocyte supplementation in Ag-specific T-B cocultures (A) Flow cytometric dot plots show the live B cells (CD19 + ) in B alone (−) and B cells supplemented with SCGF (100 ng/mL), BAFF (10 ng/mL), BAFF + SCGF, and monocytes (M) after 6 days. B + M was used as the control for 100% survivability. (B) Count of live B cells in conditions mentioned in (A). (C) Line plot shows the percent survival of B cells (relative to B + M) in supplemented culture of B cells. (D) Flow cytometric contour plots show the analysis of plasma cells in T + B (−) and T + B supplemented with BAFF, BAFF + SCGF, and monocytes (M) in 9 days cocultures. (E) Plasma cell count in the conditions mentioned in (D). (F) Flow cytometric contour plots show the analysis of activated (ICOS + PD-1 + ) CD4 + T cells in the conditions mentioned in (D). (G) Count of activated T cells in the conditions mentioned in (D). (H and I) Dot plots show the background subtracted (H) plasma cell count and (I) activated T cell count in supplemented cocultures. (J) Flow cytometric contour plots show the AIM + (OX40 + CD25 + ) CD4 + T cells in T + B (−) or supplemented cocultures after 2 days. (K) Dot plots show the background subtracted AIM + CD4 + T cells. Data are represented as mean ± SEM, with each dot representing one donor. Data represent the pool of two independent experiments. Statistics are as follows: (B, C, H, I, and K) one-way ANOVA followed by Bonferroni’s multiple comparisons test and (E and G) two-tailed paired t test. See also Figures S9 and .

Article Snippet: In some B cell survival assay, B cells were supplemented with either the recombinant BAFF (#2149-BF, R&D Systems) or SCGF (#1904-SC, R&D Systems) at various concentrations.

Techniques: Control, Clinical Proteomics, Cell Counting, Two Tailed Test

Journal: Cell Reports Methods

Article Title: An efficient immunoassay for the B cell help function of SARS-CoV-2-specific memory CD4 + T cells

doi: 10.1016/j.crmeth.2022.100224

Figure Lengend Snippet:

Article Snippet: In some B cell survival assay, B cells were supplemented with either the recombinant BAFF (#2149-BF, R&D Systems) or SCGF (#1904-SC, R&D Systems) at various concentrations.

Techniques: Purification, Recombinant, Staining, Enzyme-linked Immunosorbent Assay, Enzyme-linked Immunospot, Software

Journal: Cell stem cell

Article Title: TFAP2C- and p63-Dependent Networks Sequentially Rearrange Chromatin Landscapes to Drive Human Epidermal Lineage Commitment

doi: 10.1016/j.stem.2018.12.012

Figure Lengend Snippet: KEY RESOURCES TABLE

Article Snippet: Primary antibodies are: K18 (1:800, R&D AF7619), K14 (1:800, BioLegend SIG-3476–100); AP-2γ (1:100, Cell Signaling 2320), p63 (1:100 Gene Tex GTX102425), ITGA6 (1:200, Millipore, MAB1378).

Techniques: Purification, Recombinant, Knock-Out, Blocking Assay, Plasmid Preparation, Cloning, One Step RT-PCR, SYBR Green Assay, Magnetic Beads, Staining, Sequencing, CRISPR, Negative Control, shRNA, Expressing, Software

Journal: PLoS ONE

Article Title: Rapid and Sensitive Lentivirus Vector-Based Conditional Gene Expression Assay to Monitor and Quantify Cell Fusion Activity

doi: 10.1371/journal.pone.0010954

Figure Lengend Snippet: ( A ) Luminometric analysis of cell lysates generated from co-cultures initiated with 10 5 myoblast FLPe and 10 5 myoblast GS.Luc cells and exposed for 3 days to differentiation medium (white bar), to differentiation medium supplemented with 0.1, 0.5 and 2.5 µM SB 203580 (black bars) or to differentiation medium containing a final concentration of vehicle equivalent to that applied to co-cultures incubated with 2.5 µM SB 203580 (grey bar). Cumulative data are presented as means ± standard deviations ( n = 3). RLU, relative light units. ( B ) Western blot analysis of p38α levels in protein lysates of parental myoblasts and myoblasts GS.Luc (mock) and of myoblasts and myoblasts GS.Luc stably transduced with shRNA modules designed to down-regulate expression of eGFP (sh.eGFP), hif1α (sh.hif1α) and human p38α (sh.p38α.35, sh.p38α.33, sh.p38α.36 and sh.p38α.34). The α- and β-tubulins served as loading control. ( C ) Diagram outlining the experimental set-up applied to investigate the impact of post-transcriptional down-regulation of p38α expression on human myocyte fusion (see text for details). ( D ) Quantification through chemiluminescence of myoblast fusion activity in co-cultures consisting of a 1∶1 mixture of FLPe- and GS.Luc-encoding myoblasts either not transduced (none) or stably transduced with shRNAs sh.p38α.33, sh.p38α.36 or sh.hif1α. Data were derived from a minimum of 3 and a maximum of 6 different experiments and are presented as means ± standard error of the mean. RLU, relative light units.

Article Snippet: After incubation with blocking solution (10 mM Tris-HCl [pH 8.0], 150 mM NaCl and 0.05% Tween-20 [TBST]) supplemented with 10% (w/v) non-fat dry milk powder (Elk, Campina), the membranes were incubated overnight at 4°C with the rabbit anti-human p38α affinity-purified IgG antibody AF8691 (R&D systems) diluted 1∶3000 in blocking solution.

Techniques: Generated, Concentration Assay, Incubation, Western Blot, Stable Transfection, Transduction, shRNA, Expressing, Control, Activity Assay, Derivative Assay