amyloid oligomers (a11) antibody  (StressMarq)

Journal: bioRxiv

Article Title: c-MYC is an aggregation-prone, amyloidogenic protein

doi: 10.64898/2026.03.12.711438

Figure Lengend Snippet: (A) Detection of detergent-soluble and - insoluble c-MYC proteins by immunoblotting in HeLa cells pre-treated with 30 μM CR, followed by HS at 43°C for 30 min (representative images of three independent experiments). (B) and (C) Immunoprecipitation of either exogenously expressed HA-c-MYC or endogenous c-MYC proteins in HeLa cells by A11 antibodies (representative images of three independent experiments). (D) Detection of endogenous c-MYC proteins in HeLa cells by PLA using both the goat anti-c-MYC Ab and the rabbit anti-AO (A11) Ab. Nuclei are counterstained with Hoechst 33342. Scale bar: 10 μm. (E)-(G) Detection of endogenous c-MYC proteins in human prostate adenocarcinoma, hepatocellular carcinoma, and Alzheimer’s disease brain tissues by brightfield PLA using both the goat anti-c-MYC Ab and the rabbit anti-AO (A11) Ab. Scale bar: 100 μm for low magnification and 10 μm for high magnification. (H): Quantitation of endogenous c-MYC recognized by both the goat anti-c-MYC Ab and the rabbit anti-AO (A11) Ab through In-Cell PLA ELISA in HeLa cells treated with CR (mean ± SD, n=3 independent experiments, One-way ANOVA).

Article Snippet: All antibodies were purchased commercially, including rabbit polyclonal anti-amyloid oligomer (A11) Ab (cat#SPC-506, StressMarq Biosciences Inc.), rabbit monoclonal anti-c-MYC/N-MYC (D3N8F) Ab (cat#13987, Cell Signaling Technology), rabbit monoclonal ani-MAX (E6F6Y) Ab (cat#17471, Cell Signaling Technology), mouse monoclonal anti-HSC/HSP70 (W27) Ab (cat#sc-24, Santa Cruz Biotechnology Inc.), mouse monoclonal anti-HSP90α/β (F-8) Ab (cat#sc-13119, Santa Cruz Biotechnology Inc.), rabbit monoclonal anti-cleaved caspase 3 (Asp175) (D3E9) Ab (cat#9579, Cell Signaling Technology), goat polyclonal anti-c-MYC Ab (cat#AF3696, R&D Systems Inc.), mouse monoclonal anti-HA Ab (cat#901513, BioLegend Inc.), mouse monoclonal anti-βActin (GT5512) Ab (cat#GTX629630, GeneTex Inc.), normal rabbit IgG (cat#02-6102,ThermoFisher Scientific Inc.), Peroxidase AffiniPure Goat Anti-Rabbit IgG (H+L) (cat#111-035-144, Jackson ImmunoResearch Inc.), Peroxidase AffiniPure Goat Anti-Mouse IgG (H+L) (cat#115-035-003, Jackson ImmunoResearch Inc.), and Peroxidase IgG Fraction Monoclonal Mouse Anti-Goat IgG, light chain specific (cat#205-032-176, Jackson ImmunoResearch Inc.).

Techniques: Western Blot, Immunoprecipitation, Quantitation Assay, Enzyme-linked Immunosorbent Assay

Journal: bioRxiv

Article Title: c-MYC is an aggregation-prone, amyloidogenic protein

doi: 10.64898/2026.03.12.711438

Figure Lengend Snippet: (A) In vitro fibrillation assays using recombinant c-MYC proteins (mean ± SEM, n=3 independent experiments, Two-way ANOVA). (B) ELISA quantitation of A11 + –AOs generated by recombinant c-MYC proteins in vitro (mean ± SD, n=3 independent experiments, One-way ANOVA). (C) TEM visualization of protofibrils formed by recombinant c-MYC proteins in vitro (representative images of three independent experiments). Scale bars: 100 nm. (D) In vitro fibrillation assays using recombinant c-MYC/MAX dimers (mean ± SEM, n=3 independent experiments, Two-way ANOVA). (E) TEM visualization of recombinant c-MYC/MAX dimers in vitro (representative images of three independent experiments). Scale bars: 100 nm. (F) Quantitation of endogenous c-MYC AOs in HeLa cells treated with 10058-F4 by In-Cell PLA ELISA (mean ± SD, n=3 independent experiments, One-way ANOVA).

Article Snippet: All antibodies were purchased commercially, including rabbit polyclonal anti-amyloid oligomer (A11) Ab (cat#SPC-506, StressMarq Biosciences Inc.), rabbit monoclonal anti-c-MYC/N-MYC (D3N8F) Ab (cat#13987, Cell Signaling Technology), rabbit monoclonal ani-MAX (E6F6Y) Ab (cat#17471, Cell Signaling Technology), mouse monoclonal anti-HSC/HSP70 (W27) Ab (cat#sc-24, Santa Cruz Biotechnology Inc.), mouse monoclonal anti-HSP90α/β (F-8) Ab (cat#sc-13119, Santa Cruz Biotechnology Inc.), rabbit monoclonal anti-cleaved caspase 3 (Asp175) (D3E9) Ab (cat#9579, Cell Signaling Technology), goat polyclonal anti-c-MYC Ab (cat#AF3696, R&D Systems Inc.), mouse monoclonal anti-HA Ab (cat#901513, BioLegend Inc.), mouse monoclonal anti-βActin (GT5512) Ab (cat#GTX629630, GeneTex Inc.), normal rabbit IgG (cat#02-6102,ThermoFisher Scientific Inc.), Peroxidase AffiniPure Goat Anti-Rabbit IgG (H+L) (cat#111-035-144, Jackson ImmunoResearch Inc.), Peroxidase AffiniPure Goat Anti-Mouse IgG (H+L) (cat#115-035-003, Jackson ImmunoResearch Inc.), and Peroxidase IgG Fraction Monoclonal Mouse Anti-Goat IgG, light chain specific (cat#205-032-176, Jackson ImmunoResearch Inc.).

Techniques: In Vitro, Recombinant, Enzyme-linked Immunosorbent Assay, Quantitation Assay, Generated

Journal: bioRxiv

Article Title: c-MYC is an aggregation-prone, amyloidogenic protein

doi: 10.64898/2026.03.12.711438

Figure Lengend Snippet: (A) In vitro fibrillation assays using the human c-MYC peptide library (mean ± SEM, n=3 independent experiments, Two-way ANOVA). (B) ELISA quantitation of A11 + –AOs generated by synthetic c-MYC peptides. Results of three independent experiments are presented as a heatmap. FC: fold change. (C) and (D) In vitro fibrillation assays using the P2 and P12 peptides that are subjected to alanine scan. Results are presented as heatmaps using the averages of three independent experiments.

Article Snippet: All antibodies were purchased commercially, including rabbit polyclonal anti-amyloid oligomer (A11) Ab (cat#SPC-506, StressMarq Biosciences Inc.), rabbit monoclonal anti-c-MYC/N-MYC (D3N8F) Ab (cat#13987, Cell Signaling Technology), rabbit monoclonal ani-MAX (E6F6Y) Ab (cat#17471, Cell Signaling Technology), mouse monoclonal anti-HSC/HSP70 (W27) Ab (cat#sc-24, Santa Cruz Biotechnology Inc.), mouse monoclonal anti-HSP90α/β (F-8) Ab (cat#sc-13119, Santa Cruz Biotechnology Inc.), rabbit monoclonal anti-cleaved caspase 3 (Asp175) (D3E9) Ab (cat#9579, Cell Signaling Technology), goat polyclonal anti-c-MYC Ab (cat#AF3696, R&D Systems Inc.), mouse monoclonal anti-HA Ab (cat#901513, BioLegend Inc.), mouse monoclonal anti-βActin (GT5512) Ab (cat#GTX629630, GeneTex Inc.), normal rabbit IgG (cat#02-6102,ThermoFisher Scientific Inc.), Peroxidase AffiniPure Goat Anti-Rabbit IgG (H+L) (cat#111-035-144, Jackson ImmunoResearch Inc.), Peroxidase AffiniPure Goat Anti-Mouse IgG (H+L) (cat#115-035-003, Jackson ImmunoResearch Inc.), and Peroxidase IgG Fraction Monoclonal Mouse Anti-Goat IgG, light chain specific (cat#205-032-176, Jackson ImmunoResearch Inc.).

Techniques: In Vitro, Enzyme-linked Immunosorbent Assay, Quantitation Assay, Generated

Journal: Bioelectronic Medicine

Article Title: Electrical stimulation of the vagus nerve improves amyloid pathology in delirium superimposed on dementia

doi: 10.1186/s42234-025-00194-5

Figure Lengend Snippet: Cytokine activation of bEnd.3 monolayers drive microglia-mediated assembly of amyloid in transwell co-culture model of BBB-microglia crosstalk. A Schematic illustration of transwell model system and associated treatments and assays. Monolayers of bEnd.3 cells were formed on transwell inserts and assembled over BV-2 cells cultured on coverslips with a 3 h acclimation period in reduced serum conditions. Co-cultures were treated overnight with a combination of Cytomix with Aβ42 monomers and corresponding vehicle control combinations (as outlined in Table ). Changes in barrier health were assayed via TEER B with complementary immunocytochemical analysis of microglial activation and assembly of Aβ42 into higher-order assemblies (C-E). (B) TEER measurements of barrier integrity and activation are represented as % change compared to steady-state monolayer values. C Representative immunofluorescence images of BV-2 cells with or without Aβ42 monomers after bEnd.3 treatment with or without Cytomix. ΙΒΑ−1 (blue), 6E10 (green), OC (cyan), A11 (magenta). Scale bar 15 μm. D Quantification of microgliosis based on IBA-1 reactivity. E - H Quantification of Aβ42 species based on total amyloid content (6E10 reactivity) and respective unaggregated (6E10 solo reactivity), fibrillar (6E10 OC double reactivity), and oligomeric (6E10 A11 double reactivity) fractions. I Representative images and quantification of Aβ (magenta), Iba1 (green), and Vcam1 (red) immunoreactivity in the hippocampus of 5xFAD mice 24 h after orthopedic surgery vs. controls. Scale bar: 200 μm and 20 μm (high mag). J - L Quantification of Aβ, Iba1 and Vcam1 presented as percentage of the area. Data presented as mean ± SEM and all quantifications analyzed for statistical significance using a two-way ANOVA on a full-effects model and Fisher’s LSD. *p < 0.05; **p < 0.01; ***p < 0.001; n = 3/treatment or unpaired t-test (J-L). **p < 0.01; n = 6/group. Abbreviations: Veh: vehicle, C: control, Sx: tibia surgery

Article Snippet: Rbt , A11 Oligomer , 1:200 , StressMarq , SPC-506 , Gt , AF568 , 1:750 , Thermo , A-11,036.

Techniques: Activation Assay, Co-Culture Assay, Cell Culture, Control, Immunofluorescence

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: bioRxiv

Article Title: Driving proteomic imbalance to combat Neurofibromatosis type I (NF1)-associated malignancy

doi: 10.1101/2024.05.24.595838

Figure Lengend Snippet: (A) Violin plots of HSF1 expression in human cancer tissues showing NF1 deletion (median and IQR, n=6424, 1598, or 88, Kruskal-Wallis test). Data are generated by the TGCA Research Network ( https://www.cancer.gov/tcga ). (B) Kaplan-Meier survival curves of patients whose tumors harbor NF1 deletion (n=420 or 419, Log-rank test). Data are generated by the TGCA Research Network ( https://www.cancer.gov/tcga ). (C) and (D) The impacts of HSF1 depletion on protein polyubiquitination in both 90-8TL cells and primary human Schwann cells. Cells were transduced with lentiviral shRNAs for 4 days and global protein Lys48-specific ubiquitination was detected by immunoblotting in both detergent-soluble and -insoluble fractions of whole cell lysates (C). Protein ubiquitination was quantitated using Fiji imaging software (D) (mean ± SD, n=3 independent experiments, Two-way ANOVA). (E) and (F) Congo Red (CR) staining of 90-8TL cells and primary human Schwann cells with and without HSF1 depletion, analyzed by flow cytometry (E). Quantitation of CR staining using the median fluorescence intensity (F) (mean ± SD, n=3 independent experiments, One-way ANOVA). (G)-(J) Quantitation of soluble amyloid oligomers and insoluble amyloid fibrils by ELISA using A11 and OC antibodies, respectively (mean ± SD, n=3 independent experiments, One-way ANOVA).

Article Snippet: Following permeabilization using 0.3% Triton-X-100, samples were incubated with primary antibodies A11-FITC (1:100 in 5% BSA, StressMarq Biosciences, cat# SPC-506D-FITC) or OC-PerCP (1:100 in 5% BSA, StressMarq Biosciences, cat# SPC-507D-PCP) at RT for 3 hrs.

Techniques: Expressing, Generated, Transduction, Western Blot, Imaging, Software, Staining, Flow Cytometry, Quantitation Assay, Fluorescence, Enzyme-linked Immunosorbent Assay

Journal: bioRxiv

Article Title: Driving proteomic imbalance to combat Neurofibromatosis type I (NF1)-associated malignancy

doi: 10.1101/2024.05.24.595838

Figure Lengend Snippet: (A) Immunoblotting detection of JNK activation, mTORC1 activity, and protein polyubiquitination in S462 cells following stable TSC2 knockdown and/or HSF1 inhibition by 10 μM DTHIB for 3 days. (B) Quantitation of soluble AOs by flow cytometry using A11 antibody staining in S462 cells with and without TSC2 knockdown and 10 μM DTHIB treatment (mean ± SD, n=3 independent experiments, Two-way ANOVA). (C) Quantitation of cytotoxicity by flow cytometry using Live-or-Dye stains in S462 cells with and without stable TSC2 knockdown and DTHIB (10 μM) or combined DTHIB and LY2584702 (20 μM) treatment (mean ± SD, n=3 independent experiments, Two-way ANOVA). Cells were pre-treated with LY2584702 for 1 day followed by DTHIB treatment for another 3 days. (D) Quantitation of cytotoxicity by flow cytometry using Live-or-Dye stains in S462 cells with and without stable TSC2 knockdown and DTHIB (10 μM) or combined DTHIB and CR (30 μM) treatment (mean ± SD, n=3 independent experiments, Two-way ANOVA). Cells were pre-treated with CR for 1 day followed by DTHIB treatment for another 3 days. (E) Immunoblotting detection of mTORC1 stimulation by 500 μM NV-5138 in S462 cells with and without 10 μM DTHIB treatment. (F) Quantitation of soluble AOs by flow cytometry using A11 antibody staining in S462 cells with and without 500 μM NV-5138 stimulation and 10 μM DTHIB treatment (mean ± SD, n=3 independent experiments, One-way ANOVA). (G) Quantitation of cytotoxicity by flow cytometry using Live-or-Dye stains in S462 cells with and without 500 μM NV-5138 stimulation and 10 μM DTHIB or combined DTHIB and CR treatment (mean ± SD, n=5 independent experiments, One-way ANOVA). (H) Quantitation of cytotoxicity by flow cytometry using Live-or-Dye stains in immortalized human Schwann cells with and without 500 μM NV-5138 stimulation and 10 μM DTHIB treatment (mean ± SD, n=3 independent experiments, One-way ANOVA). (I) Immunoblotting detection of mTORC1 stimulation by 500 μM NV-5138 in immortalized human Schwann cells with and without 10 μM DTHIB treatment.

Article Snippet: Following permeabilization using 0.3% Triton-X-100, samples were incubated with primary antibodies A11-FITC (1:100 in 5% BSA, StressMarq Biosciences, cat# SPC-506D-FITC) or OC-PerCP (1:100 in 5% BSA, StressMarq Biosciences, cat# SPC-507D-PCP) at RT for 3 hrs.

Techniques: Western Blot, Activation Assay, Activity Assay, Knockdown, Inhibition, Quantitation Assay, Flow Cytometry, Staining