goat polyclonal  (R&D Systems)

Journal: Cellular & Molecular Biology Letters

Article Title: Unveiling the hidden role of the interaction between CD36 and FcγRIIb: implications for autoimmune disorders

doi: 10.1186/s11658-024-00593-7

Figure Lengend Snippet: Mice that lack CD36-expressing B cells exhibit compromised autoimmunity. A The immunization strategy employed for the Mb1 cre and Cd36 fl/fl Mb1 cre mice. Mice were immunized four times weekly with apoptotic cells (4xAC), while PBS was used as the negative control. B The gating strategy and frequency of germinal center (GC) B cells were examined. C Measurement of anti-DNA IgG levels via ELISA. The data are representative of three independent experiments. * P < 0.05, ** P < 0.01 and *** P < 0.001 (Mann–Whitney)

Article Snippet: To conduct immunoprecipitation, a total of 2 µg of goat anti-mouse CD36 antibody (R&D) or normal goat IgG (Millipore) was incubated with Pierce™ Protein A/G Magnetic Beads (Thermo Scientific™) overnight.

Techniques: Expressing, Negative Control, Enzyme-linked Immunosorbent Assay, MANN-WHITNEY

Journal: Cellular & Molecular Biology Letters

Article Title: Unveiling the hidden role of the interaction between CD36 and FcγRIIb: implications for autoimmune disorders

doi: 10.1186/s11658-024-00593-7

Figure Lengend Snippet: CD36-associated proteins were identified through co-immunoprecipitation followed by mass spectrometry analysis. A The pseudocolor plot illustrates the staining of CD36 in B cells that were either stimulated with CPG (CPG Stim) or unstimulated (Uns). B Confocal microscopy was used to analyze CD36 staining in B cells treated with CPG. C Immunoblot analysis of soluble proteins derived from B cells stimulated with CPG was subsequently performed using an anti-CD36 antibody. The total B cell protein served as the loading (positive) control referred to as Input (Inp). The B cell protein was immunoprecipitated using isotype (negative) control IgG antibodies (IgG) or anti-CD36 antibodies (CD36). D Evidence of the CD36 protein network predicted by the STRING, demonstrating its functional association with the top ten proteins. Network nodes represent proteins. Colored nodes represent the query protein and its first shell of interactors, while white nodes represent the query protein and the second shell of interactors. Filled nodes or empty nodes indicate predicted or unpredicted 3D structures, and edges represent protein–protein associations. Line color indicates the different types of interaction evidence. E Confidence view of the CD36 protein network. The line of thickness visually depicts stronger associations. F , G Gene Ontology enrichment analysis was also conducted to investigate the biological processes and molecular functions associated with the proteins identified through anti-CD36 immunoprecipitation. The data are representative of three independent experiments. * P < 0.05, ** P < 0.01 and *** P < 0.001 (Mann–Whitney)

Article Snippet: To conduct immunoprecipitation, a total of 2 µg of goat anti-mouse CD36 antibody (R&D) or normal goat IgG (Millipore) was incubated with Pierce™ Protein A/G Magnetic Beads (Thermo Scientific™) overnight.

Techniques: Immunoprecipitation, Mass Spectrometry, Staining, Confocal Microscopy, Western Blot, Derivative Assay, Positive Control, Negative Control, Functional Assay, MANN-WHITNEY

Journal: Cellular & Molecular Biology Letters

Article Title: Unveiling the hidden role of the interaction between CD36 and FcγRIIb: implications for autoimmune disorders

doi: 10.1186/s11658-024-00593-7

Figure Lengend Snippet: The protein interaction between CD36 and FcγRIIb was confirmed through various experimental approaches. A CD36 and FcγRIIb staining analysis of CPG-stimulated B cells using confocal microscopy. B The proteins immunoprecipitated by anti-CD36 antibodies were detected using anti-FcγRIIb antibodies in B cells stimulated with cytidine phosphate guanosine (CPG) or lipopolysaccharide (LPS). The total B cell protein served as the loading (positive) control referred to as Input (Inp). The B cell protein was immunoprecipitated using isotype (negative) control IgG antibodies (IgG) or anti-CD36 antibodies (CD36). C , D Docking scores and ligand mean square deviation (rmsd) for the models of CD36 docking with FcγRIIb by HDOCK SERVER. The X-axis represents the rank of the top 10 models, and the Y-axis shows the docking energy scores in C . The ligand rmsd is calculated by comparing the ligands in the CD36 model with the modeled FcγRIIb structures in D . E , F The front and back sides of the model 1 of CD36 (multicolor) docking with FcγRIIb (purple)

Article Snippet: To conduct immunoprecipitation, a total of 2 µg of goat anti-mouse CD36 antibody (R&D) or normal goat IgG (Millipore) was incubated with Pierce™ Protein A/G Magnetic Beads (Thermo Scientific™) overnight.

Techniques: Staining, Confocal Microscopy, Immunoprecipitation, Positive Control, Negative Control

Journal: Cellular & Molecular Biology Letters

Article Title: Unveiling the hidden role of the interaction between CD36 and FcγRIIb: implications for autoimmune disorders

doi: 10.1186/s11658-024-00593-7

Figure Lengend Snippet: Complex template information for CD36 and FcγRIIb

Article Snippet: To conduct immunoprecipitation, a total of 2 µg of goat anti-mouse CD36 antibody (R&D) or normal goat IgG (Millipore) was incubated with Pierce™ Protein A/G Magnetic Beads (Thermo Scientific™) overnight.

Techniques:

Journal: Cellular & Molecular Biology Letters

Article Title: Unveiling the hidden role of the interaction between CD36 and FcγRIIb: implications for autoimmune disorders

doi: 10.1186/s11658-024-00593-7

Figure Lengend Snippet: Interface residue pairs and ligand root mean square deviation (rmsd) (Å) for Model 1 of CD36 docking with FcγRIIb

Article Snippet: To conduct immunoprecipitation, a total of 2 µg of goat anti-mouse CD36 antibody (R&D) or normal goat IgG (Millipore) was incubated with Pierce™ Protein A/G Magnetic Beads (Thermo Scientific™) overnight.

Techniques: Residue

Journal: Cellular & Molecular Biology Letters

Article Title: Unveiling the hidden role of the interaction between CD36 and FcγRIIb: implications for autoimmune disorders

doi: 10.1186/s11658-024-00593-7

Figure Lengend Snippet: Loss of FcγRIIb in mice downregulates CD36 expression in B cells. A The gating strategy and scatter plot depict CD36 staining in marginal zone B cells (MZB) (CD19 + CD21 + CD23 mid ) from both wild-type (WT) and Fcgr2b knockout ( Fcgr2b KO) mice determined via flow cytometry. B The immunization strategy involved apoptotic cells (4xAC) in WT and Fcgr2b KO mice. PBS served as the negative control. C The CD36 staining was performed on MZB in WT and Fcgr2b KO mice under injections. D , E The gating strategy and scatter plot illustrate the CD36 staining pattern in germinal center B cells (GC) (B220 + IgD − CD95 + GL-7 + ) of and plasma cells (PC) (B220 int CD138 + ) in immunized WT and Fcgr2b KO mice. F The FcγRIIb staining on B cells, follicular B cells (FOB), and marginal zone B cells (MZB). G The immunization strategy was used for both WT and Cd36 knockout ( Cd36 KO) mice with four times weekly apoptotic cells (4xAC) or PBS. H The FcγRIIb staining of MZB in WT and Cd36 KO mice after treatments. The data are representative of three independent experiments. * P < 0.05, ** P < 0.01 and *** P < 0.001 (Mann–Whitney)

Article Snippet: To conduct immunoprecipitation, a total of 2 µg of goat anti-mouse CD36 antibody (R&D) or normal goat IgG (Millipore) was incubated with Pierce™ Protein A/G Magnetic Beads (Thermo Scientific™) overnight.

Techniques: Expressing, Staining, Knock-Out, Flow Cytometry, Negative Control, MANN-WHITNEY

Journal: Biochemistry and Biophysics Reports

Article Title: Nalmefene, an opioid receptor modulator, aggravates atherosclerotic plaque formation in apolipoprotein E knockout mice by enhancing oxidized low-density lipoprotein uptake in macrophages

doi: 10.1016/j.bbrep.2024.101688

Figure Lengend Snippet: Effect of nalmefene on the protein expression levels of CD36 in RAW264.7 cells. Cells were treated with nalmefene (0–300 μg/mL) for 24 h. (A) Representative immunoblots showing the protein expression levels of CD36. (B) Quantitative analysis of CD36 expression GAPDH was used as a loading control. Each dot shows data obtained from individual experiment. Data represent means ± SD (n = 4) * P < 0.05, *** P < 0.001.

Article Snippet: The blot was incubated with goat anti-CD36 (1:500; R&D Systems, Minneapolis, MN, USA) and mouse anti-glyceraldehyde 3-phosphate dehydrogenase (GAPDH; 1:2000; Millipore) antibodies at 4 °C overnight, and then incubated with horseradish peroxidase-conjugated anti-goat and -mouse (1:10000 dilution, Bio-Rad Laboratories, Hercules, CA, USA) antibodies for 60 min at room temperature.

Techniques: Expressing, Western Blot

Journal: Plants

Article Title: Botanical Antcin K Alleviates High-Fat Damage in Palm Acid Oil-Treated Vascular Endothelial Cells and Macrophages

doi: 10.3390/plants11212812

Figure Lengend Snippet: Antcin K treatments effectively reduced the expression of CD36 in palm oil-treated vascular endothelial cells. ( A ) Immunofluorescence staining to examine expressions of CD36 (green) and nuclei (blue, DAPI) in vascular endothelial cells with and without palm acid oil (PA) treatments, and with and without antcin K treatments. ( B ) Comparison of the quantified expression of CD36 of vascular endothelial cells with and without 0.75 mM palm acid oil (PA) treatment, and with and without 20 μg/mL antcin K treatment ( n = 3 for each group; values are presented as mean ± SEM, * p < 0.05, ** p < 0.01, one-way ANOVA followed by the Student–Newman–Keuls multiple comparison post hoc test).

Article Snippet: After washing three times with PBS, the tissues or cells were added 2% BSA to dilute the primary antibody of human anti-mouse VCAM-1 (1:200; sc-13160; Santa Cruz Biotechnology Inc., Dallas, TX, USA), rabbit anti-mouse KLF4 polyclone antibody (membrane protein) (1:200; GTX101508; Genetex, Irvine, CA, USA), or goat anti-mouse CD36 monoclonal antibody (1:200; sc-7309; Santa Cruz Biotechnology Inc.), and then incubated overnight in a humidified dark box at 4 °C.

Techniques: Expressing, Immunofluorescence, Staining, Comparison

Journal: Plants

Article Title: Botanical Antcin K Alleviates High-Fat Damage in Palm Acid Oil-Treated Vascular Endothelial Cells and Macrophages

doi: 10.3390/plants11212812

Figure Lengend Snippet: Possible therapeutic mechanisms of antcin K in alleviating the high-fat damage of vascular endothelial cells and macrophages in blood vessels. Antcin K treatment can (1) decrease the expression of VCAM-1 that alleviates circulating monocytes adhering to endothelial cells and migrating into subendothelial space; (2) reduce ROS generation and oxidative stress that alleviate the oxidative modification of lipoproteins and phospholipids; (3) enhance the expression of KLF4 in macrophages and endothelial cells that decrease lipid deposition and prevent macrophages converting into foam cells; (4) decrease the content of TNF-α and IL-1β; and (4) decrease the expression of CD36 that slows transformation of macrophages and endothelial cells to foam cells.

Article Snippet: After washing three times with PBS, the tissues or cells were added 2% BSA to dilute the primary antibody of human anti-mouse VCAM-1 (1:200; sc-13160; Santa Cruz Biotechnology Inc., Dallas, TX, USA), rabbit anti-mouse KLF4 polyclone antibody (membrane protein) (1:200; GTX101508; Genetex, Irvine, CA, USA), or goat anti-mouse CD36 monoclonal antibody (1:200; sc-7309; Santa Cruz Biotechnology Inc.), and then incubated overnight in a humidified dark box at 4 °C.

Techniques: Expressing, Modification, Transformation Assay

Journal: Frontiers in Cell and Developmental Biology

Article Title: Malaria Parasites Hijack Host Receptors From Exosomes to Capture Lipoproteins

doi: 10.3389/fcell.2021.749153

Figure Lengend Snippet: HDL-uptake by Plasmodium -infected erythrocytes. (A) Uptake of HDL components by P. b erghei-pRBCs. Phospholipid and ester components were concomitantly observed around parasite, but HDL was not taken up by the nRBCs (arrows). Scale bar: 5 μm. (B) Pb -pRBCs western blotted for Apo A-I, using a polyclonal primary Ab reactive to both mouse and human Apo A-I. The membrane fraction showed Apo A-I positivity with or without the addition of human HDL. (C) Flow cytometric analysis of DiO-labelled HDL uptake. Synchronized ring-stage P. falciparum incubated for 24 h with 500 μg/ml of HDL. HDL uptake was competitively inhibited by unlabeled HDL. (D) Chemical structure of BLT-1. (E) BLT-1 dose-dependent inhibition of DiO-labelled HDL uptake to Pb -pRBCs. The numbers in panels represent fraction (%) of DiO-positive cells out of total cells.

Article Snippet: Exosomes from 50 μl of equally pooled plasma from five mice per group were sequentially incubated at room temperature for 1 h with goat polyclonal anti-CD36 Ab (AF2519, R and D Systems) and HRP-labelled bovine anti-goat IgG heavy and light chains, known to be minimally cross-reactive with mouse and rat serum proteins (805–035-180, Jackson ImmunoResearch, West Grove, PA, United States).

Techniques: Infection, Western Blot, Membrane, Incubation, Inhibition

Journal: Frontiers in Cell and Developmental Biology

Article Title: Malaria Parasites Hijack Host Receptors From Exosomes to Capture Lipoproteins

doi: 10.3389/fcell.2021.749153

Figure Lengend Snippet: Immunocytochemical analysis of Pb -pRBCs for scavenger receptors and CD41. (A) Immunocytochemical analysis of Pb -pRBCs. The CD36 signals (pink) were weak in the pRBCs on days 5 and 8 (ring stage), but were abundantly detected on days 11 and 14 (schizonts). On day 11, CD36 appeared as a spot corresponding to each nucleus in the schizont’s body. SR-B1, which was detected in the pRBC membrane, was negligible in the internal parasites (arrow). Scale bar: 5 μm (B) Immunocytochemical analysis of CD36 in BM-transplanted mouse erythrocytes 6 days after Pb infection. CD36 (pink) was detected in pRBCs from the Kusabira-Orange mouse, but not in the BM-transplanted or CD36 −/− mouse. Scale bar: 5 μm. (C) Immunocytochemical analysis of pRBCs. CD41 (pink) signals were absent on days 5 and 8 (ring stage), but a small signal emanated from the internal parasites on day 8. CD41-specific fluorescence became abundant on days 11 and 14 (schizont stage). On day 11, CD41 appeared as spots corresponding to nuclei in the schizont’s body, as was also seen with CD36. Scale bar: 5 μm. (D) Co-localization of CD36 (red) and CD41 (green). Scale bar: 5 μm.

Article Snippet: Exosomes from 50 μl of equally pooled plasma from five mice per group were sequentially incubated at room temperature for 1 h with goat polyclonal anti-CD36 Ab (AF2519, R and D Systems) and HRP-labelled bovine anti-goat IgG heavy and light chains, known to be minimally cross-reactive with mouse and rat serum proteins (805–035-180, Jackson ImmunoResearch, West Grove, PA, United States).

Techniques: Membrane, Infection, Fluorescence

Journal: Frontiers in Cell and Developmental Biology

Article Title: Malaria Parasites Hijack Host Receptors From Exosomes to Capture Lipoproteins

doi: 10.3389/fcell.2021.749153

Figure Lengend Snippet: Analyses of the exosomes isolated from Pb -parasitized mouse plasma or from cultured cells differentiated from CMK11-5 cells. (A) Electron micrograph of the plasma exosomes (arrows) on day 8 post-infection. Scale bar: 333 nm. (B) Western blot of exosomes from plasma, platelets, and pRBCs. CD36, CD41 and an exosome marker (ALIX) were detected on days 5 and 8 post-infection. (C) Detection of exosomes doubly-positive for CD36 and CD41 by sandwich ELISA (duplicated). Each group of exosomes was isolated from the ‘pooled plasma’ of five mice. PBS and exosomes from a CD36 −/− mouse were used as negative controls. On days 5 and 8 post-infection, increased OD450 nm was observed. (D) immunoblot analysis for CD36 and CD41 in cultured cells differentiated from CMK11-5 cells or the exosomes released from those cells. (E) Immunocytochemical analysis of erythrocytes from a Pb -parasitized CD36 −/− mouse 6 h after adding CMK11-5-derived exosomes. CD36 and CD41 were detected from internal Pb (pink). NC: parasitized erythrocytes incubated with CMK11-5-derived exosomes and immunostained without primary Abs but with secondary Abs. Scale bar: 5 μm. (F) Trafficking of DiI-labelled exosomes derived from CMK11-5 cells into erythrocytes 1 h after exosome addition. DiI was observed in pRBCs (red) but not in uninfected ones. Scale bar: 5 μm.

Article Snippet: Exosomes from 50 μl of equally pooled plasma from five mice per group were sequentially incubated at room temperature for 1 h with goat polyclonal anti-CD36 Ab (AF2519, R and D Systems) and HRP-labelled bovine anti-goat IgG heavy and light chains, known to be minimally cross-reactive with mouse and rat serum proteins (805–035-180, Jackson ImmunoResearch, West Grove, PA, United States).

Techniques: Isolation, Clinical Proteomics, Cell Culture, Infection, Western Blot, Marker, Sandwich ELISA, Derivative Assay, Incubation

Journal: Frontiers in Cell and Developmental Biology

Article Title: Malaria Parasites Hijack Host Receptors From Exosomes to Capture Lipoproteins

doi: 10.3389/fcell.2021.749153

Figure Lengend Snippet: In vivo Pb survival and growth (parasitemia). (A) Kaplan-Meier survival curve of Pb -infected mice. CD36 −/− mice survived for significantly longer than the C57BL6 mice. (B) Parasitemia during Pb infection.

Article Snippet: Exosomes from 50 μl of equally pooled plasma from five mice per group were sequentially incubated at room temperature for 1 h with goat polyclonal anti-CD36 Ab (AF2519, R and D Systems) and HRP-labelled bovine anti-goat IgG heavy and light chains, known to be minimally cross-reactive with mouse and rat serum proteins (805–035-180, Jackson ImmunoResearch, West Grove, PA, United States).

Techniques: In Vivo, Infection

Journal: Frontiers in Cell and Developmental Biology

Article Title: Malaria Parasites Hijack Host Receptors From Exosomes to Capture Lipoproteins

doi: 10.3389/fcell.2021.749153

Figure Lengend Snippet: Proposed mechanism of exosome-based CD36 delivery and HDL capture in pRBCs. Pf EMP1 in P. falciparum and proteins that have similar binding ability to CD36 in mice malaria catch exosomes to hijack the receptors.

Article Snippet: Exosomes from 50 μl of equally pooled plasma from five mice per group were sequentially incubated at room temperature for 1 h with goat polyclonal anti-CD36 Ab (AF2519, R and D Systems) and HRP-labelled bovine anti-goat IgG heavy and light chains, known to be minimally cross-reactive with mouse and rat serum proteins (805–035-180, Jackson ImmunoResearch, West Grove, PA, United States).

Techniques: Binding Assay