goat anti- myd88  (Absolute Biotech Inc)

Journal: Frontiers in Immunology

Article Title: Dual roles of in situ generated HSP70 in antigen delivery and immunoregulation

doi: 10.3389/fimmu.2025.1638948

Figure Lengend Snippet: RFA induces TIRAP/MyD88 but not TRAM/TRIF association. (A–D) . Mice was subjected to RFA or Sham treatment or ID injection of 5 µg LPS. Skin was collected 6 h later and subjected to cryo-sectioning and PLA analysis of close association of MyD88 and TIRAP (A, B) as well as TRIF and TRAM (C, D) . Representative PLA images were shown in (A, C) and quantitative results were shown in (B, D) . Arrows point to PLA signals. E. Skin sections in RFA groups in the above studies were also stained with fluorescence-conjugated anti-MHC II antibodies. Z-stack pictures were captured and used to create 3D images. Representative 3D pictures showing the overlapping of RFA-induced TIRAP/MyD88 PLA signals with MHC II. Arrows point to overlapping signals (yellow). (F) Pie chart of PLA signals overlapped with MHC II (blue). Scale: 100 µm in (A, C, E) . One-way ANOVA with Newman-Keuls multiple comparison test was used to compare differences between groups in (B, D) . n=14–20 in (B) and n=6–8 in (D) . Over 50 PLA signals were explored by investigators in (F) . *, p<0.05; **, p<0.01; ***, p<0.001. Data are representative of three independent experiments with similar results.

Article Snippet: Goat polyclonal antibody against mouse/rat MyD88 (AF3109) was purchased from R & D Systems (Minneapolis, MN).

Techniques: Injection, Staining, Fluorescence, Comparison

Journal: Frontiers in Immunology

Article Title: Dual roles of in situ generated HSP70 in antigen delivery and immunoregulation

doi: 10.3389/fimmu.2025.1638948

Figure Lengend Snippet: HSP70 suppresses RFA-induced TLR4/IRAK/NFκB signaling. (A, B) WT and HSP70 KO mice were subjected to RFA or Sham treatment or ID injection of LPS or PBS. Skin was collected 6 h later in (A, B) IP and IB were conducted to evaluate TLR4/TIRAP binding (A) and IRAK4/IRAK1 binding in (B, C) . WT, HSP70 KO, MyD88 KO, TLR2 KO, and TLR4 KO mice were subjected to RFA or Sham treatment or ID injection of LPS. Skin was collected 2 h later. Cytosol and nuclear fractions were separated and nuclear fraction was analyzed by WB analysis to detect phosphorylated p65 using Lamin b1 as a loading control. (D) Skin IL-6 levels 6 h after RFA, Sham, or LPS treatment of lateral back skin of WT, HSP70 KO, TLR2 KO, TLR4 KO, and MyD88 KO mice. Two-way ANOVA with Fisher’s LSD test was used to compare differences between groups. n=4-6. *, p<0.05; **, p<0.01; ***, p<0.001. Original membrane pictures were shown in Supplementary Figure S9 . Data are representative of two independent experiments with similar results.

Article Snippet: Goat polyclonal antibody against mouse/rat MyD88 (AF3109) was purchased from R & D Systems (Minneapolis, MN).

Techniques: Injection, Binding Assay, Control, Membrane

Journal: Biomolecules

Article Title: Novel Inhibitory Actions of Neuroactive Steroid [3α,5α]-3-Hydroxypregnan-20-One on Toll-like Receptor 4-Dependent Neuroimmune Signaling

doi: 10.3390/biom14111441

Figure Lengend Snippet: 3α,5α-THP inhibits TIRAP binding to MyD88 in P rat male ( A ) and female ( B ) hippocampi. MyD88 immunoprecipitation of various components of the myddosome complex was conducted in male and female, vehicle- (45% w / v 2-hydroxypropyl-β-cyclodextrin; 30 min) and 3α,5α-THP-treated (15 mg/kg; 30 min) P rats ( A , B ). Densiometric comparison of the effect of 3α,5α-THP on MyD88 immunoprecipitation of TIRAP ( A , B ), IRAK4 ( C , D ) and IRAK1 ( E , F ) in female and male hippocampi. Immunoblots and densiometric measurements of extracted hippocampal TIRAP ( G ) and MyD88 ( H ) in vehicle- and 3α,5α-THP-treated animals. Western blot original images are in the . *** p < 0.005, **** p < 0.001, ns: no significant difference.

Article Snippet: CelLytic extracted protein lysates (500 μg) were incubated with goat anti-MyD88 antibody (6 μg: R&D Systems Cat. #AF3109) and incubated for 4 h (4 °C).

Techniques: Binding Assay, Immunoprecipitation, Comparison, Western Blot

Journal: CNS Neuroscience & Therapeutics

Article Title: Intracerebral hemorrhage‐induced brain injury in mice: The role of peroxiredoxin 2‐Toll ‐like receptor 4 inflammatory axis

doi: 10.1111/cns.14681

Figure Lengend Snippet: TLR (toll‐like receptor) signaling pathway is upregulated in the inflammation induced by Prx2 (peroxiredoxin 2). (A) KEGG (Kyoto Encyclopedia of Genes and Genomes) pathway analysis of DEGs (differentially expressed genes) in the ipsilateral basal ganglia at 24 h after saline or Prx2 injection (10 μL); n = 5 for saline and n = 7 for Prx2. Dot color represents Q‐value from the least significant (green) to the most significant (red); dot size represents gene number, the number of significant DEGs in the KEGG pathway; rich factor represents the fraction of significant DEGs among all genes in the KEGG pathway. (B) FPKM (Mean fragments per kilobase of transcript per million mapped reads) values of TLR2, TLR4, Myd88 (myeloid differentiation primary response protein 88) in the ipsilateral basal ganglia at 24 h after saline or Prx2 injection. Values are mean ± SD; n = 5 for saline and n = 7 for Prx2; # p < 0.01 versus saline group. (C) TLR4 immunofluorescence at the ipsilateral basal ganglia 24 h after saline or Prx2 injection. Values are mean ± SD; n = 7 for both groups; # p < 0.01 versus saline group. Scale bar = 200 μm at low magnification, 20 μm at high magnification.

Article Snippet: The primary antibodies used for immunohistochemical staining and immunofluorescence staining were rabbit anti‐Prx2 IgG (10545‐2‐AP, 1:200; Proteintech, Wuhan, China), rabbit anti‐IBA‐1 (ionized calcium‐binding adaptor molecule 1, 10904‐1‐AP, 1:400; Proteintech), rabbit anti‐MPO IgG (myeloperoxidase, PA5‐16672, 1:400; ThermoFisher Scientific), and goat anti‐MyD88 (myeloid differentiation primary response protein 88, EB06667, 1:200; Everest Biotech, Oxford, UK).

Techniques: Saline, Injection, Immunofluorescence

Journal: CNS Neuroscience & Therapeutics

Article Title: Intracerebral hemorrhage‐induced brain injury in mice: The role of peroxiredoxin 2‐Toll ‐like receptor 4 inflammatory axis

doi: 10.1111/cns.14681

Figure Lengend Snippet: Intracaudate injection of Prx2 (peroxiredoxin 2) activates TLR4 (toll‐like receptor 4) signaling pathway. (A) Triple immunofluorescence labeling of IBA‐1 (ionized calcium‐binding adaptor molecule 1) and TLR4 at the ipsilateral basal ganglia 24 h after Prx2 injection. Co‐localization of TLR4 with IBA‐1 (microglia/macrophage marker). Scale bar = 200 μm at low magnification, 20 μm at high magnification, 10 μm at the highest magnification. (B) Triple immunofluorescence labeling of MPO (myeloperoxidase) and TLR4 at the ipsilateral basal ganglia 24 h after Prx2 injection. Co‐localization of TLR4 with MPO (neutrophil marker). Scale bar = 200 μm at low magnification, 20 μm at high magnification, 5 μm at the highest magnification. (C) Triple immunofluorescence labeling of TLR4 and Myd88 (myeloid differentiation primary response protein 88) at the ipsilateral basal ganglia 24 h after saline or Prx2 injection. Values are mean ± SD; n = 7 for both groups; # p < 0.01 versus saline group. Scale bar = 200 μm at low magnification, 20 μm at high magnification.

Article Snippet: The primary antibodies used for immunohistochemical staining and immunofluorescence staining were rabbit anti‐Prx2 IgG (10545‐2‐AP, 1:200; Proteintech, Wuhan, China), rabbit anti‐IBA‐1 (ionized calcium‐binding adaptor molecule 1, 10904‐1‐AP, 1:400; Proteintech), rabbit anti‐MPO IgG (myeloperoxidase, PA5‐16672, 1:400; ThermoFisher Scientific), and goat anti‐MyD88 (myeloid differentiation primary response protein 88, EB06667, 1:200; Everest Biotech, Oxford, UK).

Techniques: Injection, Immunofluorescence, Labeling, Binding Assay, Marker, Saline