Journal: bioRxiv

Article Title: The β-amyloid oligomer Aβ*56 is associated with Alzheimer’s dementia independently of amyloid pathology

doi: 10.1101/2025.04.11.648457

Figure Lengend Snippet: ( a, c ) Schematic diagrams illustrating IP/WB settings for detecting Aβ(40)*56 (a) and Aβ(42)*56 (c). hAPP, human amyloid precursor protein; Aβ, β-amyloid. ( b, d ) Upper , representative IP/WB showing that D8Q71 (b)- or D3E10 (d)-precipitated proteins from aqueous brain extracts contain ∼56-kDa entities (arrow) and monomeric Aβ (#) that are detected by biotinylated 82E1 (82E1-biotin); middle , WB showing NeuN proteins (arrowheads); bottom , WB showing GAPDH proteins (@). NCI, no cognitive impairment; MCI, mild cognitive impairment; AD, Alzheimer’s disease dementia. + and -, IP reactions containing a capture antibody and aqueous brain extracts of APP/TTA mice (+) or littermates expressing no hAPP (-) (Online Resource 1 : Table S4). ⁋, IP reactions containing only the capture antibodies. †, IP reactions containing a generic rabbit IgG and aqueous brain extracts (combined from the six individuals shown in b or d, each contributing equivalent protein masses). ‡, synthetic Aβ(1-40) (b) or Aβ(1-42) (d). ( e ) A schematic illustration of isolating Aβ*56 variants from human brains. ( f, g ) Representative IP/WB showing that the detection of D8Q7I (f)- or D3E10 (g)-purified ∼56-kDa entities by anti-oligomer A11 antibodies following treatment with Tris buffer but not 6 M GuHCl. ( h ) Representative WB showing that ∼56-kDa entities (arrow, fractions [Frac.] 51-61 and 63-71), ∼14-kDa entities (%, Frac. 65-79), ∼9-kDa entities ($, Frac. 67-77), and monomeric Aβ (#, Frac. 35-45 and 75-83) are detected in SEC fractions of aqueous AD brain extracts using 82E1-biotin. Lower panels, longer exposure. Std., biomolecule standards. ( i ) Quantification of levels of the ∼56-kDa entities and monomeric Aβ shown in (h). The level of the ∼56-kDa entities in each fraction is normalized to the highest level in fraction 57 and of monomeric Aβ (Aβ mono.), fraction 37. kDa, kilo-Dalton. *, non-specific bands.

Article Snippet: For WB analyses using biotinylated A11 (StressMarq Biosciences, Victoria, BC; Cat. No.: SPC-506D-BI; RRID: AB_10962958) antibodies ( Fig. 1 f and ) and their associated secondary detection agent horseradish peroxidase (HRP)-conjugated NeutrAvidin (NA) (Online Resource 1 : Figure S2), we used blocking buffer 1 (10 mM Tris-HCl, pH 7.4; 200 mM NaCl; 0.01% (v/v) polyoxyethylene (20) sorbitan monolaurate (Tween 20; MilliporeSigma, Cat. No.: P9416-100ML); and 5% (w/v) Carnation instant non-fat dry milk powder (Nestlé, Rosslyn, VA; Cat. No.: 12428935)); and for all other WB analyses, we used blocking buffer 2 (10 mM Tris-HCl, pH 7.4; 200 mM NaCl; 0.1% (v/v) Tween 20; and 5% (w/v) bovine serum albumin (MilliporeSigma, Cat. No.: A3803-100G)).

Techniques: Expressing, Purification

Journal: iScience

Article Title: Natural killer cells and dendritic epidermal γδ T cells orchestrate type 1 conventional DC spatiotemporal repositioning toward CD8 + T cells

doi: 10.1016/j.isci.2021.103059

Figure Lengend Snippet:

Article Snippet: Anti Fluorescein/Oregon Green A488 (Polyclonal) , ThermoFisher , #A11090.

Techniques: Purification, Avidin-Biotin Assay, Virus, Recombinant, High Molecular Weight, Cell Isolation, Microarray, Software

Journal: Frontiers in Immunology

Article Title: An orally available cancer drug AZD6738 prevents type 1 diabetes

doi: 10.3389/fimmu.2023.1290058

Figure Lengend Snippet: ATRi prevents type 1 diabetes by inhibiting proliferation of diabetogenic CD4 + T cells. (A) In vivo ATRi treatment schedule. NOD.scid mice were adoptively transferred with 2 million activated CD4 + T cells isolated from NOD.BDC2.5 mice. Mice were treated with vehicle (black) or indicated doses of ATRi by oral gavage (og) for 1 (red), 2 (blue), or 5 weeks (green). Type 1 diabetes incident (B) as well as % body weight change (C) of these mice were monitored. After 5 weeks of treatment, splenocytes from 2-week (blue, diabetic) or 5-week (green, diabetes free) ATRi treated splenocytes were harvested and transferred into a new set of NOD.scid mice. (D) T1D incident of newly transferred mice. (E, F) flow cytometry of transferred splenocytes. (E) population of CD4 + T cells within the splenocytes (in live gate). (F) % effector memory CD4 + T cells (left) and representative CD62L versus CD44 plot of 2-week or 5-week ATRi treated splenocytes (within live/CD4 + gate). T N : naïve T cells, T CM : central memory T cells, T EM : effector memory T cells. (G-J) Splenocytes from NOD.BDC2.5 mice were activated ex vivo with 0.25 μg/ml BDC2.5 mimotope and treated with vehicle (black) or 5 μM ATRi (red). (G-J) ATRi was given at the same time as activation. Survival (G) , effector function as indicated by IFNγ levels (H) , as well as proliferation indicated by the dilution of cell cycle violet (CTV) dye (I) at 24-, 48-, and 72-hours post activation are shown. Increased CTV dye dilution (decreased CTV intensity) represents increased proliferation. (J) Cells were treated for 4 hours starting at 48 hours post stimulation and DNA damage indicated by γH2AX levels are shown. (B-F) n=5, not significant (no asterisk), **: p<0.01, ***: p<0.001, ****: p<0.0001 by Mentel-Cox test (B, D) , Tukey (C) , or unpaired T-test (E, F) . (G, I) n=6 (three biological replicates with two technical replicate each). (H) n=10-12 (two biological replicates with six technical replicate each). (J) n=6 (two biological replicates with three technical replicates each). (G-J) not significant (no asterisk), **: p<0.01, ***: p<0.001, ****: p<0.0001 by unpaired T-test.

Article Snippet: Anti-mouse CD28 (BD 553294), CD3 3 (Fisher BDB553058), CD4 PE (Fisher BDB553730) at 1:250, CD8 APC (Fisher BDB553035) at 1:250, CD44 BV785 (BioLegend 103041) at 1:500, CD62L PE-Cy7 (BioLegend 104418) at 1:250, TCRβ A488 (BioLegend 109215) at 1:250, γH2AX A488 (BioLegend 613404) at 1:250, IFNγ (BD 551216), biotin IFNγ (BD 554410).

Techniques: In Vivo, Isolation, Flow Cytometry, Ex Vivo, Activation Assay