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pcdna3 1 human lrrc33  (Addgene inc)


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    Structured Review

    Addgene inc pcdna3 1 human lrrc33
    A Immunoblot comparing lysates of ipsilateral cortices from wild-type and Rubcn-mutant mice probed with Rubicon A (RbA) antibody. B Western blot showing immunoprecipitation of Rubicon protein from brain lysates of wild-type and Rubcn-mutant mice using Rubicon A (RbA) and Rubicon B (RbA) antibodies. * represents background signal. Blot is probed with RbB antibody. C Rubicon protein (amino-acid) sequence highlighting peptides (p1-p12 in blue) used for peptide mapping of wild-type and mutant RUBCN proteins. Red circle is highlighting Met296 as a potential alternative translation initiation site. D Normalized peptide abundance across the length of RUBCN protein in wild-type and Rubcn-mutant immunoprecipitates using RbA and RbB antibodies. E Comparative enrichment analysis showing fold change of proteins enriched in wild-type over Rubcn-mutant immunoprecipitates using RbA and RbB antibodies. F Immunoprecipitation assay showing lysates of HEK293T cells transfected with indicated plasmids for 24 h and immunoprecipitated with anti-FLAG or anti-HA agarose beads. Left: Immunoblot of immunoprecipitates with the indicated antibodies. Right: Fold change of <t>NRROS</t> signal in EGFP-Rubicon-Flag and Nrros-Myc/His (Nrros+Rubicon1-3) co-transfected cells to EGFP-C1 and Nrros-Myc/His (Nrros ONLY) co-transfected cells. Each FLAG IP sample was normalized to corresponding HA IP sample for quantification. Sample size (n) = 3 biological replicates for EGFP-Rubicon-Flag and Nrros-Myc/His co-transfected cells. G Immunoblot of whole cell lysates used for immunoprecipitation assay in F. H Immunoprecipitation assay showing lysates of HEK293T cells transfected with indicated plasmids for 24 h and immunoprecipitated with anti-MYC or anti-IgG antibodies. Left: Immunoblot of immunoprecipitates with the indicated antibodies. Right: Fold change of RUBCN signal in EGFP-Rubicon-Flag and Nrros-Myc/His (Nrros+Rubicon1-2) co-transfected cells to EGFP-C1 and EGFP-Rubicon-Flag (Rubicon ONLY) co-transfected cells. Each MYC IP sample was normalized to corresponding IgG IP sample. Sample size (n) = 2 biological replicates for EGFP-Rubicon-Flag and Nrros-Myc/His co-transfected cells. I Immunoblot of whole cell lysates used for immunoprecipitation assay in H.
    Pcdna3 1 Human Lrrc33, supplied by Addgene inc, used in various techniques. Bioz Stars score: 94/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/product/egfp+rubicon+%CE%B4ct/EGFP-Rubicon+deltaCT+(Plasmid+%2328040)/bio_rxiv__64898__2026__03__04__709622-216-17-19
    Average 94 stars, based on 1 article reviews
    pcdna3 1 human lrrc33 - by Bioz Stars, 2026-09
    94/100 stars

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    1) Product Images from "Rubicon modulates neuroimmune responses following traumatic brain injury"

    Article Title: Rubicon modulates neuroimmune responses following traumatic brain injury

    Journal: bioRxiv

    doi: 10.64898/2026.03.04.709622

    A Immunoblot comparing lysates of ipsilateral cortices from wild-type and Rubcn-mutant mice probed with Rubicon A (RbA) antibody. B Western blot showing immunoprecipitation of Rubicon protein from brain lysates of wild-type and Rubcn-mutant mice using Rubicon A (RbA) and Rubicon B (RbA) antibodies. * represents background signal. Blot is probed with RbB antibody. C Rubicon protein (amino-acid) sequence highlighting peptides (p1-p12 in blue) used for peptide mapping of wild-type and mutant RUBCN proteins. Red circle is highlighting Met296 as a potential alternative translation initiation site. D Normalized peptide abundance across the length of RUBCN protein in wild-type and Rubcn-mutant immunoprecipitates using RbA and RbB antibodies. E Comparative enrichment analysis showing fold change of proteins enriched in wild-type over Rubcn-mutant immunoprecipitates using RbA and RbB antibodies. F Immunoprecipitation assay showing lysates of HEK293T cells transfected with indicated plasmids for 24 h and immunoprecipitated with anti-FLAG or anti-HA agarose beads. Left: Immunoblot of immunoprecipitates with the indicated antibodies. Right: Fold change of NRROS signal in EGFP-Rubicon-Flag and Nrros-Myc/His (Nrros+Rubicon1-3) co-transfected cells to EGFP-C1 and Nrros-Myc/His (Nrros ONLY) co-transfected cells. Each FLAG IP sample was normalized to corresponding HA IP sample for quantification. Sample size (n) = 3 biological replicates for EGFP-Rubicon-Flag and Nrros-Myc/His co-transfected cells. G Immunoblot of whole cell lysates used for immunoprecipitation assay in F. H Immunoprecipitation assay showing lysates of HEK293T cells transfected with indicated plasmids for 24 h and immunoprecipitated with anti-MYC or anti-IgG antibodies. Left: Immunoblot of immunoprecipitates with the indicated antibodies. Right: Fold change of RUBCN signal in EGFP-Rubicon-Flag and Nrros-Myc/His (Nrros+Rubicon1-2) co-transfected cells to EGFP-C1 and EGFP-Rubicon-Flag (Rubicon ONLY) co-transfected cells. Each MYC IP sample was normalized to corresponding IgG IP sample. Sample size (n) = 2 biological replicates for EGFP-Rubicon-Flag and Nrros-Myc/His co-transfected cells. I Immunoblot of whole cell lysates used for immunoprecipitation assay in H.
    Figure Legend Snippet: A Immunoblot comparing lysates of ipsilateral cortices from wild-type and Rubcn-mutant mice probed with Rubicon A (RbA) antibody. B Western blot showing immunoprecipitation of Rubicon protein from brain lysates of wild-type and Rubcn-mutant mice using Rubicon A (RbA) and Rubicon B (RbA) antibodies. * represents background signal. Blot is probed with RbB antibody. C Rubicon protein (amino-acid) sequence highlighting peptides (p1-p12 in blue) used for peptide mapping of wild-type and mutant RUBCN proteins. Red circle is highlighting Met296 as a potential alternative translation initiation site. D Normalized peptide abundance across the length of RUBCN protein in wild-type and Rubcn-mutant immunoprecipitates using RbA and RbB antibodies. E Comparative enrichment analysis showing fold change of proteins enriched in wild-type over Rubcn-mutant immunoprecipitates using RbA and RbB antibodies. F Immunoprecipitation assay showing lysates of HEK293T cells transfected with indicated plasmids for 24 h and immunoprecipitated with anti-FLAG or anti-HA agarose beads. Left: Immunoblot of immunoprecipitates with the indicated antibodies. Right: Fold change of NRROS signal in EGFP-Rubicon-Flag and Nrros-Myc/His (Nrros+Rubicon1-3) co-transfected cells to EGFP-C1 and Nrros-Myc/His (Nrros ONLY) co-transfected cells. Each FLAG IP sample was normalized to corresponding HA IP sample for quantification. Sample size (n) = 3 biological replicates for EGFP-Rubicon-Flag and Nrros-Myc/His co-transfected cells. G Immunoblot of whole cell lysates used for immunoprecipitation assay in F. H Immunoprecipitation assay showing lysates of HEK293T cells transfected with indicated plasmids for 24 h and immunoprecipitated with anti-MYC or anti-IgG antibodies. Left: Immunoblot of immunoprecipitates with the indicated antibodies. Right: Fold change of RUBCN signal in EGFP-Rubicon-Flag and Nrros-Myc/His (Nrros+Rubicon1-2) co-transfected cells to EGFP-C1 and EGFP-Rubicon-Flag (Rubicon ONLY) co-transfected cells. Each MYC IP sample was normalized to corresponding IgG IP sample. Sample size (n) = 2 biological replicates for EGFP-Rubicon-Flag and Nrros-Myc/His co-transfected cells. I Immunoblot of whole cell lysates used for immunoprecipitation assay in H.

    Techniques Used: Western Blot, Mutagenesis, Immunoprecipitation, Sequencing, Transfection

    Related Articles

    Transfection:

    Article Title: Rubicon modulates neuroimmune responses following traumatic brain injury
    Article Snippet: HEK293T cells were maintained in Dulbecco’s Modified Eagle Medium (DMEM) (Gibco; 11995073) supplemented with 10% fetal bovine serum (FBS) (GeminiBio; 900-108H-500) and 1% antibiotic-antimycotic (Gibco; 15240062). .. HEK293T cells were transiently transfected with EGFP-Rubicon (Addgene; 28022), EGFP-Rubicon Δ182 (Addgene; 28041), EGFP-Rubicon ΔCT (Addgene; 28040), pcDNA3.1(+)-human LRRC33 (Addgene; 111602), and/or pEGFP-C1 vectors using Lipofectamine TM 3000 Transfection Reagent (ThermoFisher Scientific; L3000015) for 24 h.( , , ) The cell monolayers were washed with ice-cold PBS and lysed in cell lysis bucer (50 mM Tris-HCl, 150 mM NaCl, 1 mM EDTA, and 1% Triton X-100 or 0.5% Igepal CA-630 (SigmaAldrich; I8896) containing protease and phosphatase inhibitors. .. The cell lysates were centrifuged at 18,000 xg for 30 min, and the resulting supernatants were pre-cleared with Pierce TM Protein A/G Agarose beads (ThermoFisher Scientific; 20421) and mouse mAb IgG1 (CST; 5415) or mouse IgG2a (CST; 61656) for 2 h at 4 °C with rotation.

    Lysis:

    Article Title: Rubicon modulates neuroimmune responses following traumatic brain injury
    Article Snippet: HEK293T cells were maintained in Dulbecco’s Modified Eagle Medium (DMEM) (Gibco; 11995073) supplemented with 10% fetal bovine serum (FBS) (GeminiBio; 900-108H-500) and 1% antibiotic-antimycotic (Gibco; 15240062). .. HEK293T cells were transiently transfected with EGFP-Rubicon (Addgene; 28022), EGFP-Rubicon Δ182 (Addgene; 28041), EGFP-Rubicon ΔCT (Addgene; 28040), pcDNA3.1(+)-human LRRC33 (Addgene; 111602), and/or pEGFP-C1 vectors using Lipofectamine TM 3000 Transfection Reagent (ThermoFisher Scientific; L3000015) for 24 h.( , , ) The cell monolayers were washed with ice-cold PBS and lysed in cell lysis bucer (50 mM Tris-HCl, 150 mM NaCl, 1 mM EDTA, and 1% Triton X-100 or 0.5% Igepal CA-630 (SigmaAldrich; I8896) containing protease and phosphatase inhibitors. .. The cell lysates were centrifuged at 18,000 xg for 30 min, and the resulting supernatants were pre-cleared with Pierce TM Protein A/G Agarose beads (ThermoFisher Scientific; 20421) and mouse mAb IgG1 (CST; 5415) or mouse IgG2a (CST; 61656) for 2 h at 4 °C with rotation.



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    pcdna3 1 human lrrc33  (Addgene inc)
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    Addgene inc pcdna3 1 human lrrc33
    A Immunoblot comparing lysates of ipsilateral cortices from wild-type and Rubcn-mutant mice probed with Rubicon A (RbA) antibody. B Western blot showing immunoprecipitation of Rubicon protein from brain lysates of wild-type and Rubcn-mutant mice using Rubicon A (RbA) and Rubicon B (RbA) antibodies. * represents background signal. Blot is probed with RbB antibody. C Rubicon protein (amino-acid) sequence highlighting peptides (p1-p12 in blue) used for peptide mapping of wild-type and mutant RUBCN proteins. Red circle is highlighting Met296 as a potential alternative translation initiation site. D Normalized peptide abundance across the length of RUBCN protein in wild-type and Rubcn-mutant immunoprecipitates using RbA and RbB antibodies. E Comparative enrichment analysis showing fold change of proteins enriched in wild-type over Rubcn-mutant immunoprecipitates using RbA and RbB antibodies. F Immunoprecipitation assay showing lysates of HEK293T cells transfected with indicated plasmids for 24 h and immunoprecipitated with anti-FLAG or anti-HA agarose beads. Left: Immunoblot of immunoprecipitates with the indicated antibodies. Right: Fold change of <t>NRROS</t> signal in EGFP-Rubicon-Flag and Nrros-Myc/His (Nrros+Rubicon1-3) co-transfected cells to EGFP-C1 and Nrros-Myc/His (Nrros ONLY) co-transfected cells. Each FLAG IP sample was normalized to corresponding HA IP sample for quantification. Sample size (n) = 3 biological replicates for EGFP-Rubicon-Flag and Nrros-Myc/His co-transfected cells. G Immunoblot of whole cell lysates used for immunoprecipitation assay in F. H Immunoprecipitation assay showing lysates of HEK293T cells transfected with indicated plasmids for 24 h and immunoprecipitated with anti-MYC or anti-IgG antibodies. Left: Immunoblot of immunoprecipitates with the indicated antibodies. Right: Fold change of RUBCN signal in EGFP-Rubicon-Flag and Nrros-Myc/His (Nrros+Rubicon1-2) co-transfected cells to EGFP-C1 and EGFP-Rubicon-Flag (Rubicon ONLY) co-transfected cells. Each MYC IP sample was normalized to corresponding IgG IP sample. Sample size (n) = 2 biological replicates for EGFP-Rubicon-Flag and Nrros-Myc/His co-transfected cells. I Immunoblot of whole cell lysates used for immunoprecipitation assay in H.
    Pcdna3 1 Human Lrrc33, supplied by Addgene inc, used in various techniques. Bioz Stars score: 94/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/product/egfp+rubicon+%CE%B4ct/EGFP-Rubicon+deltaCT+(Plasmid+%2328040)/bio_rxiv__64898__2026__03__04__709622-216-17-19
    Average 94 stars, based on 1 article reviews
    pcdna3 1 human lrrc33 - by Bioz Stars, 2026-09
    94/100 stars
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    egfp rubicon δct  (Addgene inc)
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    Addgene inc egfp rubicon δct
    A Immunoblot comparing lysates of ipsilateral cortices from wild-type and Rubcn-mutant mice probed with Rubicon A (RbA) antibody. B Western blot showing immunoprecipitation of Rubicon protein from brain lysates of wild-type and Rubcn-mutant mice using Rubicon A (RbA) and Rubicon B (RbA) antibodies. * represents background signal. Blot is probed with RbB antibody. C Rubicon protein (amino-acid) sequence highlighting peptides (p1-p12 in blue) used for peptide mapping of wild-type and mutant RUBCN proteins. Red circle is highlighting Met296 as a potential alternative translation initiation site. D Normalized peptide abundance across the length of RUBCN protein in wild-type and Rubcn-mutant immunoprecipitates using RbA and RbB antibodies. E Comparative enrichment analysis showing fold change of proteins enriched in wild-type over Rubcn-mutant immunoprecipitates using RbA and RbB antibodies. F Immunoprecipitation assay showing lysates of HEK293T cells transfected with indicated plasmids for 24 h and immunoprecipitated with anti-FLAG or anti-HA agarose beads. Left: Immunoblot of immunoprecipitates with the indicated antibodies. Right: Fold change of NRROS signal in <t>EGFP-Rubicon-Flag</t> and Nrros-Myc/His (Nrros+Rubicon1-3) co-transfected cells to EGFP-C1 and Nrros-Myc/His (Nrros ONLY) co-transfected cells. Each FLAG IP sample was normalized to corresponding HA IP sample for quantification. Sample size (n) = 3 biological replicates for EGFP-Rubicon-Flag and Nrros-Myc/His co-transfected cells. G Immunoblot of whole cell lysates used for immunoprecipitation assay in F. H Immunoprecipitation assay showing lysates of HEK293T cells transfected with indicated plasmids for 24 h and immunoprecipitated with anti-MYC or anti-IgG antibodies. Left: Immunoblot of immunoprecipitates with the indicated antibodies. Right: Fold change of RUBCN signal in EGFP-Rubicon-Flag and Nrros-Myc/His (Nrros+Rubicon1-2) co-transfected cells to EGFP-C1 and EGFP-Rubicon-Flag (Rubicon ONLY) co-transfected cells. Each MYC IP sample was normalized to corresponding IgG IP sample. Sample size (n) = 2 biological replicates for EGFP-Rubicon-Flag and Nrros-Myc/His co-transfected cells. I Immunoblot of whole cell lysates used for immunoprecipitation assay in H.
    Egfp Rubicon δct, supplied by Addgene inc, used in various techniques. Bioz Stars score: 94/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/product/egfp+rubicon+%CE%B4ct/EGFP-Rubicon+delta182+(Plasmid+%2328041)/bio_rxiv__64898__2026__03__04__709622-216-13-15
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    Image Search Results


    A Immunoblot comparing lysates of ipsilateral cortices from wild-type and Rubcn-mutant mice probed with Rubicon A (RbA) antibody. B Western blot showing immunoprecipitation of Rubicon protein from brain lysates of wild-type and Rubcn-mutant mice using Rubicon A (RbA) and Rubicon B (RbA) antibodies. * represents background signal. Blot is probed with RbB antibody. C Rubicon protein (amino-acid) sequence highlighting peptides (p1-p12 in blue) used for peptide mapping of wild-type and mutant RUBCN proteins. Red circle is highlighting Met296 as a potential alternative translation initiation site. D Normalized peptide abundance across the length of RUBCN protein in wild-type and Rubcn-mutant immunoprecipitates using RbA and RbB antibodies. E Comparative enrichment analysis showing fold change of proteins enriched in wild-type over Rubcn-mutant immunoprecipitates using RbA and RbB antibodies. F Immunoprecipitation assay showing lysates of HEK293T cells transfected with indicated plasmids for 24 h and immunoprecipitated with anti-FLAG or anti-HA agarose beads. Left: Immunoblot of immunoprecipitates with the indicated antibodies. Right: Fold change of NRROS signal in EGFP-Rubicon-Flag and Nrros-Myc/His (Nrros+Rubicon1-3) co-transfected cells to EGFP-C1 and Nrros-Myc/His (Nrros ONLY) co-transfected cells. Each FLAG IP sample was normalized to corresponding HA IP sample for quantification. Sample size (n) = 3 biological replicates for EGFP-Rubicon-Flag and Nrros-Myc/His co-transfected cells. G Immunoblot of whole cell lysates used for immunoprecipitation assay in F. H Immunoprecipitation assay showing lysates of HEK293T cells transfected with indicated plasmids for 24 h and immunoprecipitated with anti-MYC or anti-IgG antibodies. Left: Immunoblot of immunoprecipitates with the indicated antibodies. Right: Fold change of RUBCN signal in EGFP-Rubicon-Flag and Nrros-Myc/His (Nrros+Rubicon1-2) co-transfected cells to EGFP-C1 and EGFP-Rubicon-Flag (Rubicon ONLY) co-transfected cells. Each MYC IP sample was normalized to corresponding IgG IP sample. Sample size (n) = 2 biological replicates for EGFP-Rubicon-Flag and Nrros-Myc/His co-transfected cells. I Immunoblot of whole cell lysates used for immunoprecipitation assay in H.

    Journal: bioRxiv

    Article Title: Rubicon modulates neuroimmune responses following traumatic brain injury

    doi: 10.64898/2026.03.04.709622

    Figure Lengend Snippet: A Immunoblot comparing lysates of ipsilateral cortices from wild-type and Rubcn-mutant mice probed with Rubicon A (RbA) antibody. B Western blot showing immunoprecipitation of Rubicon protein from brain lysates of wild-type and Rubcn-mutant mice using Rubicon A (RbA) and Rubicon B (RbA) antibodies. * represents background signal. Blot is probed with RbB antibody. C Rubicon protein (amino-acid) sequence highlighting peptides (p1-p12 in blue) used for peptide mapping of wild-type and mutant RUBCN proteins. Red circle is highlighting Met296 as a potential alternative translation initiation site. D Normalized peptide abundance across the length of RUBCN protein in wild-type and Rubcn-mutant immunoprecipitates using RbA and RbB antibodies. E Comparative enrichment analysis showing fold change of proteins enriched in wild-type over Rubcn-mutant immunoprecipitates using RbA and RbB antibodies. F Immunoprecipitation assay showing lysates of HEK293T cells transfected with indicated plasmids for 24 h and immunoprecipitated with anti-FLAG or anti-HA agarose beads. Left: Immunoblot of immunoprecipitates with the indicated antibodies. Right: Fold change of NRROS signal in EGFP-Rubicon-Flag and Nrros-Myc/His (Nrros+Rubicon1-3) co-transfected cells to EGFP-C1 and Nrros-Myc/His (Nrros ONLY) co-transfected cells. Each FLAG IP sample was normalized to corresponding HA IP sample for quantification. Sample size (n) = 3 biological replicates for EGFP-Rubicon-Flag and Nrros-Myc/His co-transfected cells. G Immunoblot of whole cell lysates used for immunoprecipitation assay in F. H Immunoprecipitation assay showing lysates of HEK293T cells transfected with indicated plasmids for 24 h and immunoprecipitated with anti-MYC or anti-IgG antibodies. Left: Immunoblot of immunoprecipitates with the indicated antibodies. Right: Fold change of RUBCN signal in EGFP-Rubicon-Flag and Nrros-Myc/His (Nrros+Rubicon1-2) co-transfected cells to EGFP-C1 and EGFP-Rubicon-Flag (Rubicon ONLY) co-transfected cells. Each MYC IP sample was normalized to corresponding IgG IP sample. Sample size (n) = 2 biological replicates for EGFP-Rubicon-Flag and Nrros-Myc/His co-transfected cells. I Immunoblot of whole cell lysates used for immunoprecipitation assay in H.

    Article Snippet: HEK293T cells were transiently transfected with EGFP-Rubicon (Addgene; 28022), EGFP-Rubicon Δ182 (Addgene; 28041), EGFP-Rubicon ΔCT (Addgene; 28040), pcDNA3.1(+)-human LRRC33 (Addgene; 111602), and/or pEGFP-C1 vectors using Lipofectamine TM 3000 Transfection Reagent (ThermoFisher Scientific; L3000015) for 24 h.( , , ) The cell monolayers were washed with ice-cold PBS and lysed in cell lysis bucer (50 mM Tris-HCl, 150 mM NaCl, 1 mM EDTA, and 1% Triton X-100 or 0.5% Igepal CA-630 (SigmaAldrich; I8896) containing protease and phosphatase inhibitors.

    Techniques: Western Blot, Mutagenesis, Immunoprecipitation, Sequencing, Transfection

    A Immunoblot comparing lysates of ipsilateral cortices from wild-type and Rubcn-mutant mice probed with Rubicon A (RbA) antibody. B Western blot showing immunoprecipitation of Rubicon protein from brain lysates of wild-type and Rubcn-mutant mice using Rubicon A (RbA) and Rubicon B (RbA) antibodies. * represents background signal. Blot is probed with RbB antibody. C Rubicon protein (amino-acid) sequence highlighting peptides (p1-p12 in blue) used for peptide mapping of wild-type and mutant RUBCN proteins. Red circle is highlighting Met296 as a potential alternative translation initiation site. D Normalized peptide abundance across the length of RUBCN protein in wild-type and Rubcn-mutant immunoprecipitates using RbA and RbB antibodies. E Comparative enrichment analysis showing fold change of proteins enriched in wild-type over Rubcn-mutant immunoprecipitates using RbA and RbB antibodies. F Immunoprecipitation assay showing lysates of HEK293T cells transfected with indicated plasmids for 24 h and immunoprecipitated with anti-FLAG or anti-HA agarose beads. Left: Immunoblot of immunoprecipitates with the indicated antibodies. Right: Fold change of NRROS signal in EGFP-Rubicon-Flag and Nrros-Myc/His (Nrros+Rubicon1-3) co-transfected cells to EGFP-C1 and Nrros-Myc/His (Nrros ONLY) co-transfected cells. Each FLAG IP sample was normalized to corresponding HA IP sample for quantification. Sample size (n) = 3 biological replicates for EGFP-Rubicon-Flag and Nrros-Myc/His co-transfected cells. G Immunoblot of whole cell lysates used for immunoprecipitation assay in F. H Immunoprecipitation assay showing lysates of HEK293T cells transfected with indicated plasmids for 24 h and immunoprecipitated with anti-MYC or anti-IgG antibodies. Left: Immunoblot of immunoprecipitates with the indicated antibodies. Right: Fold change of RUBCN signal in EGFP-Rubicon-Flag and Nrros-Myc/His (Nrros+Rubicon1-2) co-transfected cells to EGFP-C1 and EGFP-Rubicon-Flag (Rubicon ONLY) co-transfected cells. Each MYC IP sample was normalized to corresponding IgG IP sample. Sample size (n) = 2 biological replicates for EGFP-Rubicon-Flag and Nrros-Myc/His co-transfected cells. I Immunoblot of whole cell lysates used for immunoprecipitation assay in H.

    Journal: bioRxiv

    Article Title: Rubicon modulates neuroimmune responses following traumatic brain injury

    doi: 10.64898/2026.03.04.709622

    Figure Lengend Snippet: A Immunoblot comparing lysates of ipsilateral cortices from wild-type and Rubcn-mutant mice probed with Rubicon A (RbA) antibody. B Western blot showing immunoprecipitation of Rubicon protein from brain lysates of wild-type and Rubcn-mutant mice using Rubicon A (RbA) and Rubicon B (RbA) antibodies. * represents background signal. Blot is probed with RbB antibody. C Rubicon protein (amino-acid) sequence highlighting peptides (p1-p12 in blue) used for peptide mapping of wild-type and mutant RUBCN proteins. Red circle is highlighting Met296 as a potential alternative translation initiation site. D Normalized peptide abundance across the length of RUBCN protein in wild-type and Rubcn-mutant immunoprecipitates using RbA and RbB antibodies. E Comparative enrichment analysis showing fold change of proteins enriched in wild-type over Rubcn-mutant immunoprecipitates using RbA and RbB antibodies. F Immunoprecipitation assay showing lysates of HEK293T cells transfected with indicated plasmids for 24 h and immunoprecipitated with anti-FLAG or anti-HA agarose beads. Left: Immunoblot of immunoprecipitates with the indicated antibodies. Right: Fold change of NRROS signal in EGFP-Rubicon-Flag and Nrros-Myc/His (Nrros+Rubicon1-3) co-transfected cells to EGFP-C1 and Nrros-Myc/His (Nrros ONLY) co-transfected cells. Each FLAG IP sample was normalized to corresponding HA IP sample for quantification. Sample size (n) = 3 biological replicates for EGFP-Rubicon-Flag and Nrros-Myc/His co-transfected cells. G Immunoblot of whole cell lysates used for immunoprecipitation assay in F. H Immunoprecipitation assay showing lysates of HEK293T cells transfected with indicated plasmids for 24 h and immunoprecipitated with anti-MYC or anti-IgG antibodies. Left: Immunoblot of immunoprecipitates with the indicated antibodies. Right: Fold change of RUBCN signal in EGFP-Rubicon-Flag and Nrros-Myc/His (Nrros+Rubicon1-2) co-transfected cells to EGFP-C1 and EGFP-Rubicon-Flag (Rubicon ONLY) co-transfected cells. Each MYC IP sample was normalized to corresponding IgG IP sample. Sample size (n) = 2 biological replicates for EGFP-Rubicon-Flag and Nrros-Myc/His co-transfected cells. I Immunoblot of whole cell lysates used for immunoprecipitation assay in H.

    Article Snippet: HEK293T cells were transiently transfected with EGFP-Rubicon (Addgene; 28022), EGFP-Rubicon Δ182 (Addgene; 28041), EGFP-Rubicon ΔCT (Addgene; 28040), pcDNA3.1(+)-human LRRC33 (Addgene; 111602), and/or pEGFP-C1 vectors using Lipofectamine TM 3000 Transfection Reagent (ThermoFisher Scientific; L3000015) for 24 h.( , , ) The cell monolayers were washed with ice-cold PBS and lysed in cell lysis bucer (50 mM Tris-HCl, 150 mM NaCl, 1 mM EDTA, and 1% Triton X-100 or 0.5% Igepal CA-630 (SigmaAldrich; I8896) containing protease and phosphatase inhibitors.

    Techniques: Western Blot, Mutagenesis, Immunoprecipitation, Sequencing, Transfection