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Cell Signaling Technology Inc tlr 2 antibody

( A ) Effect of BB1 on <t>TLR-2/TLR-6</t> or TLR-2/TLR-1 heterodimer binding and signal-transduction pathway activation as determined by SEAP (secreted embryonic alkaline phosphatase) activity in TLR-1 or TLR-6 knock-out HEK-293 reporter cell lines (InvivoGen, San Diego, CA). BB1 produced a marked increase in SEAP activity in TLR-2/TLR-6 (TLR-1 -/- ), but not TLR-2/TLR-1 (TLR-6 -/- ), expressing HEK-293 cells. PAM3cBK4 (positive control) produced an increase in SEAP activity in TLR-2/TLR-1 (TLR-6 -/- ) HEK-293 cells. The siRNA knock-down of TLR-1 or TLR-6, but not TLR-2, inhibited the BB1-induced increase in occludin mRNA ( B ) and protein and densitometry analysis ( C, D ) and Caco-2 TEER ( E ) ∗∗∗∗ P < 0.0001 versus control, NS (not significant).

Tlr 2 Antibody, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 95/100, based on 216 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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1) Product Images from "Probiotic bacteria Bifidobacterium bifidum upregulation of intestinal epithelial tight junction barrier is mediated by TLR-2/TLR-6 receptor complex activation of occludin gene"

Article Title: Probiotic bacteria Bifidobacterium bifidum upregulation of intestinal epithelial tight junction barrier is mediated by TLR-2/TLR-6 receptor complex activation of occludin gene

Journal: NPJ Biofilms and Microbiomes

doi: 10.1038/s41522-025-00903-7

( A ) Effect of BB1 on TLR-2/TLR-6 or TLR-2/TLR-1 heterodimer binding and signal-transduction pathway activation as determined by SEAP (secreted embryonic alkaline phosphatase) activity in TLR-1 or TLR-6 knock-out HEK-293 reporter cell lines (InvivoGen, San Diego, CA). BB1 produced a marked increase in SEAP activity in TLR-2/TLR-6 (TLR-1 -/- ), but not TLR-2/TLR-1 (TLR-6 -/- ), expressing HEK-293 cells. PAM3cBK4 (positive control) produced an increase in SEAP activity in TLR-2/TLR-1 (TLR-6 -/- ) HEK-293 cells. The siRNA knock-down of TLR-1 or TLR-6, but not TLR-2, inhibited the BB1-induced increase in occludin mRNA ( B ) and protein and densitometry analysis ( C, D ) and Caco-2 TEER ( E ) ∗∗∗∗ P < 0.0001 versus control, NS (not significant).

Figure Legend Snippet: ( A ) Effect of BB1 on TLR-2/TLR-6 or TLR-2/TLR-1 heterodimer binding and signal-transduction pathway activation as determined by SEAP (secreted embryonic alkaline phosphatase) activity in TLR-1 or TLR-6 knock-out HEK-293 reporter cell lines (InvivoGen, San Diego, CA). BB1 produced a marked increase in SEAP activity in TLR-2/TLR-6 (TLR-1 -/- ), but not TLR-2/TLR-1 (TLR-6 -/- ), expressing HEK-293 cells. PAM3cBK4 (positive control) produced an increase in SEAP activity in TLR-2/TLR-1 (TLR-6 -/- ) HEK-293 cells. The siRNA knock-down of TLR-1 or TLR-6, but not TLR-2, inhibited the BB1-induced increase in occludin mRNA ( B ) and protein and densitometry analysis ( C, D ) and Caco-2 TEER ( E ) ∗∗∗∗ P < 0.0001 versus control, NS (not significant).

Techniques Used: Binding Assay, Transduction, Activation Assay, Activity Assay, Knock-Out, Produced, Expressing, Positive Control, Knockdown, Control

A BB1, but not BB4, treatment produced a focal aggregation of TLR-2 at the Caco-2 apical membrane surface and translocation of TOLLIP towards the apical membrane surface from the medial and basal regions of Caco-2 cells, as assessed by immunostaining. (TOLLIP, green; TLR-2 red; and DAPI blue). Scale bars = 10 μm. Quantification of TOLLIP apical translocation immunostaining intensity using ImageJ. ∗∗∗∗ P < 0.0001 versus control, NS (not significant) ( B ). The siRNA knockdown of TLR-2 or IRAK-1 inhibited the BB1-induced apical translocation of TOLLIP. The siRNA-induced of TOLLIP inhibited the BB1-induced increase in occludin mRNA ( C ) and protein expression ( D ), densitometry ( E ) and Caco-2 TEER ( F ). ∗∗∗∗ P < 0.0001 versus control, NS (not significant).

Figure Legend Snippet: A BB1, but not BB4, treatment produced a focal aggregation of TLR-2 at the Caco-2 apical membrane surface and translocation of TOLLIP towards the apical membrane surface from the medial and basal regions of Caco-2 cells, as assessed by immunostaining. (TOLLIP, green; TLR-2 red; and DAPI blue). Scale bars = 10 μm. Quantification of TOLLIP apical translocation immunostaining intensity using ImageJ. ∗∗∗∗ P < 0.0001 versus control, NS (not significant) ( B ). The siRNA knockdown of TLR-2 or IRAK-1 inhibited the BB1-induced apical translocation of TOLLIP. The siRNA-induced of TOLLIP inhibited the BB1-induced increase in occludin mRNA ( C ) and protein expression ( D ), densitometry ( E ) and Caco-2 TEER ( F ). ∗∗∗∗ P < 0.0001 versus control, NS (not significant).

Techniques Used: Produced, Membrane, Translocation Assay, Immunostaining, Control, Knockdown, Expressing

A BB1, but not BB4, caused an increase in IRAK-1 phosphorylation (Thr209) as assessed by immunoblotting. The siRNA-induced knockdown of IRAK-1 inhibited the BB1-induced increase in occludin mRNA ( B ), occludin protein ( C ), densitometry ( D ) and Caco-2 TEER ( E ) (means ± SE, n = 4). ∗∗∗∗ P < 0.0001 versus control, NS (not significant). The siRNA-induced knockdown of TLR-2 or TLR-6, but not TLR-1, inhibited the BB1 increase in IRAK-1 phosphorylation, but had no effect on total IRAK-1 protein expression ( F ).

Figure Legend Snippet: A BB1, but not BB4, caused an increase in IRAK-1 phosphorylation (Thr209) as assessed by immunoblotting. The siRNA-induced knockdown of IRAK-1 inhibited the BB1-induced increase in occludin mRNA ( B ), occludin protein ( C ), densitometry ( D ) and Caco-2 TEER ( E ) (means ± SE, n = 4). ∗∗∗∗ P < 0.0001 versus control, NS (not significant). The siRNA-induced knockdown of TLR-2 or TLR-6, but not TLR-1, inhibited the BB1 increase in IRAK-1 phosphorylation, but had no effect on total IRAK-1 protein expression ( F ).

Techniques Used: Phospho-proteomics, Western Blot, Knockdown, Control, Expressing

A Oral -gastric gavage of BB1 [1 × 10 9 colony-forming units (CFUs)/mL], but not BB4, produced a decrease in mouse small intestinal permeability to dextran 10 kDa. P **** < 0.0001 versus control. BB1 administration caused an increase in occludin mRNA ( B ) and protein ( C ), densitometry ( D ) in the mouse intestinal tissue (24-hour treatment period). E BB1 administration also produced an increase in occludin expression and localization at the apical membrane surface and junctional areas, as assess by immunostaining. (Occludin, green; nuclei, blue). Scale bars = 10 μm ∗ P < 0.05, ∗∗∗ P < 0.001, and ∗∗∗∗ P < 0.0001, NS (not significant). ( E ). BB1 oral-gastric gavage administration produced a cytoplasmic-to-apical membrane translocation of TOLLIP in mouse intestinal epithelial cells ( F ). The BB1-induced apical membrane TOLLIP recruitment was inhibited in the TLR-2 ΔIEC mice (TOLLIP, green; occludin, red; (Leica SP8 confocal microscope).

Figure Legend Snippet: A Oral -gastric gavage of BB1 [1 × 10 9 colony-forming units (CFUs)/mL], but not BB4, produced a decrease in mouse small intestinal permeability to dextran 10 kDa. P **** < 0.0001 versus control. BB1 administration caused an increase in occludin mRNA ( B ) and protein ( C ), densitometry ( D ) in the mouse intestinal tissue (24-hour treatment period). E BB1 administration also produced an increase in occludin expression and localization at the apical membrane surface and junctional areas, as assess by immunostaining. (Occludin, green; nuclei, blue). Scale bars = 10 μm ∗ P < 0.05, ∗∗∗ P < 0.001, and ∗∗∗∗ P < 0.0001, NS (not significant). ( E ). BB1 oral-gastric gavage administration produced a cytoplasmic-to-apical membrane translocation of TOLLIP in mouse intestinal epithelial cells ( F ). The BB1-induced apical membrane TOLLIP recruitment was inhibited in the TLR-2 ΔIEC mice (TOLLIP, green; occludin, red; (Leica SP8 confocal microscope).

Techniques Used: Produced, Permeability, Control, Expressing, Membrane, Immunostaining, Translocation Assay, Microscopy

A BB1 oral-gastric gavage produced an increase in small intestinal tissue expressions of TLR-2 and TLR-6 but not TLR-1. BB1 also produced an increase in apical membrane expression and localization of TLR-2 in mouse intestinal epithelial cells in the wild-type (WT) mice but not in the TLR-2 ΔIEC mice, as assessed by immunostaining (TLR-2, green; occludin, red; nuclei, blue; magnification 63x) ( B ). The BB1-induced increase in mouse intestinal tissue occludin mRNA ( C ) and protein ( D ), densitometry ( E ) and apical membrane and junctional localization of occludin (occludin, green; nuclei, blue) ( F ), and decrease in intestinal permeability to dextran 10 kDa ( G ) were inhibited in the TLR-2 ΔIEC mice. ∗∗∗∗ P < 0.0001 versus control, NS (not significant).

Figure Legend Snippet: A BB1 oral-gastric gavage produced an increase in small intestinal tissue expressions of TLR-2 and TLR-6 but not TLR-1. BB1 also produced an increase in apical membrane expression and localization of TLR-2 in mouse intestinal epithelial cells in the wild-type (WT) mice but not in the TLR-2 ΔIEC mice, as assessed by immunostaining (TLR-2, green; occludin, red; nuclei, blue; magnification 63x) ( B ). The BB1-induced increase in mouse intestinal tissue occludin mRNA ( C ) and protein ( D ), densitometry ( E ) and apical membrane and junctional localization of occludin (occludin, green; nuclei, blue) ( F ), and decrease in intestinal permeability to dextran 10 kDa ( G ) were inhibited in the TLR-2 ΔIEC mice. ∗∗∗∗ P < 0.0001 versus control, NS (not significant).

Techniques Used: Produced, Membrane, Expressing, Immunostaining, Permeability, Control

BB1 attaches to the apical membrane surface of the host intestinal epithelial cells by binding to the TLR-2/TLR-6 heterodimer complex and induces the phosphorylation of IRAK-1 and apical translocation of TOLLIP, which, in turn, leads to the down-steam activation of occludin gene and occludin protein synthesis and occludin-dependent upregulation of the intestinal TJ barrier.

Figure Legend Snippet: BB1 attaches to the apical membrane surface of the host intestinal epithelial cells by binding to the TLR-2/TLR-6 heterodimer complex and induces the phosphorylation of IRAK-1 and apical translocation of TOLLIP, which, in turn, leads to the down-steam activation of occludin gene and occludin protein synthesis and occludin-dependent upregulation of the intestinal TJ barrier.

Techniques Used: Membrane, Binding Assay, Phospho-proteomics, Translocation Assay, Serial Time-encoded Amplified Microscopy, Activation Assay

Western Blot:

Article Title: Novel Virulence Role of Pneumococcal NanA in Host Inflammation and Cell Death Through the Activation of Inflammasome and the Caspase Pathway
Article Snippet: TLR-2 , BioLegend #309702 , IP. .. TLR-2 , Cell Signaling Technology #12276S , WB. .. phospho-AMPKα (T172) , Cell Signaling Technology #4188 , WB.

Saline:

Article Title: Validation of a Quantification Method for Curcumin Derivatives and Their Hepatoprotective Effects on Nonalcoholic Fatty Liver Disease.
Article Snippet: The quantified samples were subjected to sodium dodecyl sulphate polyacrylamide gel (SDS-PAGE) electrophoresis and then transferred onto polyvinylidene difluoride (PVDF; Millipore, Bedford, MA, USA) membranes at 15 V for 45 min. .. The membranes were blocked with 3% BSA (Sigma-Aldrich, St. Louis, MO, USA) in Tris-buffered saline with Tween 20 buffer (TBST) for 2 h and were incubated with primary antibodies against ß-actin, SREBP-1c, FAS, PPAR-γ, C/EBP-α (Santa Cruz Biotechnology, Paso Robles, CA, USA), p-AMPK, AMPK, TLR-2, and TLR-4 (Cell Signaling Technology, Danvers, MA, USA) overnight at 4 ◦C. .. The membranes were then washed with TBST and incubated with anti-mouse or anti-rabbit secondary antibodies (Invitrogen, Thermo Fisher Scientific, Inc., Carlsbad, CA, USA) at room temperature.

Article Title: Validation of a Quantification Method for Curcumin Derivatives and Their Hepatoprotective Effects on Nonalcoholic Fatty Liver Disease
Article Snippet: The quantified samples were subjected to sodium dodecyl sulphate polyacrylamide gel (SDS-PAGE) electrophoresis and then transferred onto polyvinylidene difluoride (PVDF; Millipore, Bedford, MA, USA) membranes at 15 V for 45 min. .. The membranes were blocked with 3% BSA (Sigma-Aldrich, St. Louis, MO, USA) in Tris-buffered saline with Tween 20 buffer (TBST) for 2 h and were incubated with primary antibodies against ß-actin, SREBP-1c, FAS, PPAR-γ, C/EBP-α (Santa Cruz Biotechnology, Paso Robles, CA, USA), p-AMPK, AMPK, TLR-2, and TLR-4 (Cell Signaling Technology, Danvers, MA, USA) overnight at 4 °C. .. The membranes were then washed with TBST and incubated with anti-mouse or anti-rabbit secondary antibodies (Invitrogen, Thermo Fisher Scientific, Inc., Carlsbad, CA, USA) at room temperature.

Incubation:

Article Title: Corynebacterium tuberculostearicum , a human skin colonizer, induces the canonical nuclear factor‐κB inflammatory signaling pathway in human skin cells
Article Snippet: Cells were then blocked for 1 hour at room temperature in 1% BSA (catalog no. a‐4503; Sigma‐Aldrich) dissolved in PBS containing 0.01% Tween 20 (catalog no. P5927; Sigma‐Aldrich). .. Cells were subsequently incubated overnight at 4°C with antibodies against phosphorylated IκBα (mouse monoclonal antibody [catalog no. 9246; Cell Signaling]), NF‐κB‐P 65 (mouse monoclonal antibody [catalog no. SC‐293072; Santa Cruz Biotechnology]) or TLR 2 (rabbit monoclonal antibody [catalog no. 12276; Cell Signaling]) in PBS‐Tween‐BSA at the manufacturer‐recommended dilutions. .. After this incubation, cells were washed three times (5 minutes each) in PBS and incubated with Alexa Fluor 488 goat anti‐mouse secondary antibody (catalog no. A11029; Invitrogen) diluted in PBS‐Tween‐BSA (1:500) for 1 hour at room temperature, followed by three washes (5 minutes each) in PBS.

other:

Article Title: Cyanidin and delphinidin restore colon physiology in high fat diet-fed mice: Involvement of TLR-4 and redox-regulated signaling.
Article Snippet: Antibodies for IκB kinase (IKK) α (#2682), phospho (Ser176/180)IKKα/β (#2697), p65 (#8242), phospho (Ser536)-p65 (#3033), TLR-2 (#13744), phospho (Thr180/Tyr182)-p38 (#9211), extracellular signal-regulated kinase (ERK1/2) (#9102), phospho (Thr202/Tyr204)ERK1/2 (#4370), c-jun N-terminal kinase (JNK) 2 (#9258), PI3K (#5405), phospho (Ser473)-Akt (#4060), Akt (#9279), myosin light chain (MLC) (#8505), phospho (Thr18/Ser19)-MLC (#3674), MyD88 (#4283), myosin phosphatase target subunit 1 (MYPT1) (#8574), phospho (Thr696)-MYPT1 (#5163), and β-actin (#12620) were obtained from Cell Signaling Technology (Danvers, MA).

Article Title: When Two Worlds Collide: The Contribution and Association Between Genetics (APOEε4) and Neuroinflammation (IL-1β) in Alzheimer’s Neuropathogenesis
Article Snippet: TLR-2 , Rabbit mAb , #13744 , Cell Signaling Technology.

Image Search Results

a) Schematic of TLR-2 reporter allele structure and outcomes following editing. The allele includes two open reading frames encoding different fluorescent proteins; an upstream mVenus (green) in the +1 frame and downstream TagRFP (red) in the +3 frame linked by a P2A sequence. The mVenus cassette is interrupted by a 108 bp polylinker sequence that includes several guide target sequences and a stop codon (blue square). Thus in its native configuration, neither fluorescent protein is expressed. Following CRISPR/Cas9 editing, repair via NHEJ will result in expression of TagRFP if the frame shift results in a -2 deletion, or multiple thereof. Alternatively, HDR can be detected with the inclusion of a plasmid donor designed to repair the mVenus gap. b) Mouse embryo reporter validation workflow. Fertilized zygotes from TLR-2 reporter mice (typically male homozygous to WT female) are either microinjected or electroporated with Cas9 RNP with or without a plasmid donor for HDR. The embryos are then cultured to the blastocyst stage where the outcome can be scored by fluorescent imaging, and followed up by PCR-Sanger sequencing to confirm the identify of specific edits.

Journal: bioRxiv

Article Title: Novel mouse reporter models for the detection of genome editing events in vivo

doi: 10.64898/2026.04.29.721708

Figure Lengend Snippet: a) Schematic of TLR-2 reporter allele structure and outcomes following editing. The allele includes two open reading frames encoding different fluorescent proteins; an upstream mVenus (green) in the +1 frame and downstream TagRFP (red) in the +3 frame linked by a P2A sequence. The mVenus cassette is interrupted by a 108 bp polylinker sequence that includes several guide target sequences and a stop codon (blue square). Thus in its native configuration, neither fluorescent protein is expressed. Following CRISPR/Cas9 editing, repair via NHEJ will result in expression of TagRFP if the frame shift results in a -2 deletion, or multiple thereof. Alternatively, HDR can be detected with the inclusion of a plasmid donor designed to repair the mVenus gap. b) Mouse embryo reporter validation workflow. Fertilized zygotes from TLR-2 reporter mice (typically male homozygous to WT female) are either microinjected or electroporated with Cas9 RNP with or without a plasmid donor for HDR. The embryos are then cultured to the blastocyst stage where the outcome can be scored by fluorescent imaging, and followed up by PCR-Sanger sequencing to confirm the identify of specific edits.

Article Snippet: TLR-2 mice (B6.129S6- Gt(ROSA)26Sor tm 12 (CAG–Venus*,–TagRFP*) Rkuhn /MurrJ (JAXStrain # 034033; RRID:IMSR_JAX:034033) were generated with a R26-GFP_KI-TLR2 (“traffic light reporter”) targeting vector was designed to insert the TLR reporter construct (“TLR-2”) into the Gt(ROSA)26Sor locus: the TLR reporter construct (“TLR-2”) containing a CAG promoter Venus*-P2A-TagRFP-bovine hGH polyA, in a reading frame that is shifted by 2 bp (+3) .

Techniques: Sequencing, CRISPR, Expressing, Plasmid Preparation, Biomarker Discovery, Cell Culture, Imaging

a) Position of guides tested for the TLR-2 allele relative to the 108bp polylinker (gray) and TGA stop codon (blue). b) Percentage of TLR-2 heterozygous blastocysts showing visible expression of TagRFP following editing. Mouse zygotes electroporated with Cas9 RNPs with the guides listed on the X axis were cultured to the blastocysts stage and scored for expression of TagRFP, with corresponding levels of overall editing presented in (c) . Of note, the low number of blasts showing TagRFP fluorescence relative to the editing rate for R26-1 reflects its cut position 3’ to the stop codon in the polylinker. d) Percentage of TLR-2 homozyogous blastocysts with visible expression of TagRFP following editing and corresponding editing percentages shown in (e) . Data are representative of at least two technical replicates per guide and plots are showing mean and standard deviation of replicates with total number of blasts assessed across all replicates below each bar.

Journal: bioRxiv

Article Title: Novel mouse reporter models for the detection of genome editing events in vivo

doi: 10.64898/2026.04.29.721708

Figure Lengend Snippet: a) Position of guides tested for the TLR-2 allele relative to the 108bp polylinker (gray) and TGA stop codon (blue). b) Percentage of TLR-2 heterozygous blastocysts showing visible expression of TagRFP following editing. Mouse zygotes electroporated with Cas9 RNPs with the guides listed on the X axis were cultured to the blastocysts stage and scored for expression of TagRFP, with corresponding levels of overall editing presented in (c) . Of note, the low number of blasts showing TagRFP fluorescence relative to the editing rate for R26-1 reflects its cut position 3’ to the stop codon in the polylinker. d) Percentage of TLR-2 homozyogous blastocysts with visible expression of TagRFP following editing and corresponding editing percentages shown in (e) . Data are representative of at least two technical replicates per guide and plots are showing mean and standard deviation of replicates with total number of blasts assessed across all replicates below each bar.

Techniques: Expressing, Cell Culture, Fluorescence, Standard Deviation

a) To confirm the TLR-2 allele can be activated in all cells and tissues, we generated a germline model following targeting with Cas9 RNP and R26-52 guides. Founders harboring a -14 bp frameshift allele were bred to generate N1 mice. b) Imaging of 12 tissues showing widespread and consistent activation of TagRFP expression. Scale bar = 200 microns.

Journal: bioRxiv

Article Title: Novel mouse reporter models for the detection of genome editing events in vivo

doi: 10.64898/2026.04.29.721708

Figure Lengend Snippet: a) To confirm the TLR-2 allele can be activated in all cells and tissues, we generated a germline model following targeting with Cas9 RNP and R26-52 guides. Founders harboring a -14 bp frameshift allele were bred to generate N1 mice. b) Imaging of 12 tissues showing widespread and consistent activation of TagRFP expression. Scale bar = 200 microns.

Techniques: Generated, Imaging, Activation Assay, Expressing

Embryo culture testing of SauCas9 guide efficiency for both fluorescence activation (a) and editing efficiency (b) . Plots are showing mean and standard deviation of replicates with total number of blasts assessed across all replicates below each bar. SaC9-g3 showed the best performance with ∼25% fluorescence and an 80% editing rate. c) AAV9 vectors encoding SauCas9 and guide SaC9-g3 were injected IV into 10 week old adult TLR-2 mice (1 x 10 11 vg/mouse) and tissues were harvested 4 weeks later (n=8, including controls). Genotyping of the liver (d) showed up to 30% editing of the TLR-2 allele of which 1/3 were in the TagRFP frame. Mean editing and Standard Deviation is shown for the AAV9-treated animals. e) Immunofluorescence of the liver of AAV9-treated male and female mice, showing substantial RFP activation versus saline control. Scale bar = 200 microns.

Journal: bioRxiv

Article Title: Novel mouse reporter models for the detection of genome editing events in vivo

doi: 10.64898/2026.04.29.721708

Figure Lengend Snippet: Embryo culture testing of SauCas9 guide efficiency for both fluorescence activation (a) and editing efficiency (b) . Plots are showing mean and standard deviation of replicates with total number of blasts assessed across all replicates below each bar. SaC9-g3 showed the best performance with ∼25% fluorescence and an 80% editing rate. c) AAV9 vectors encoding SauCas9 and guide SaC9-g3 were injected IV into 10 week old adult TLR-2 mice (1 x 10 11 vg/mouse) and tissues were harvested 4 weeks later (n=8, including controls). Genotyping of the liver (d) showed up to 30% editing of the TLR-2 allele of which 1/3 were in the TagRFP frame. Mean editing and Standard Deviation is shown for the AAV9-treated animals. e) Immunofluorescence of the liver of AAV9-treated male and female mice, showing substantial RFP activation versus saline control. Scale bar = 200 microns.

Techniques: Embryo Culture, Fluorescence, Activation Assay, Standard Deviation, Injection, Immunofluorescence, Saline, Control

a) Schematic of the TLR-7 allele, showing the shortened polylinker sequence (30 nt) as compared to TLR-2 including a stop codon (blue), that can be repaired with an ssODN. b) Mouse blastocyst validation for NHEJ repair shows high editing rates (>95%) with an expected 1/3 of edits resulting in a shift to the +3 frame to produce the red fluorescent signal. c) Validation of HDR functionality of the allele, showing ∼30% repair indicated by mVenus fluorescence and a similar level of NHEJ activation of Katushka2S. Some blastocysts showed mosaic activation of both cassettes. Plots are showing mean and standard deviation of replicates with total number of blasts assessed across all replicates below each bar. d) Germline indel generation (2 base pair deletion) demonstrates robust, widespread Katushka2S activation in all tissues. Scale bar = 200 microns.

Journal: bioRxiv

Article Title: Novel mouse reporter models for the detection of genome editing events in vivo

doi: 10.64898/2026.04.29.721708

Figure Lengend Snippet: a) Schematic of the TLR-7 allele, showing the shortened polylinker sequence (30 nt) as compared to TLR-2 including a stop codon (blue), that can be repaired with an ssODN. b) Mouse blastocyst validation for NHEJ repair shows high editing rates (>95%) with an expected 1/3 of edits resulting in a shift to the +3 frame to produce the red fluorescent signal. c) Validation of HDR functionality of the allele, showing ∼30% repair indicated by mVenus fluorescence and a similar level of NHEJ activation of Katushka2S. Some blastocysts showed mosaic activation of both cassettes. Plots are showing mean and standard deviation of replicates with total number of blasts assessed across all replicates below each bar. d) Germline indel generation (2 base pair deletion) demonstrates robust, widespread Katushka2S activation in all tissues. Scale bar = 200 microns.

Techniques: Sequencing, Biomarker Discovery, Fluorescence, Activation Assay, Standard Deviation

Journal: NPJ Biofilms and Microbiomes

Article Title: Probiotic bacteria Bifidobacterium bifidum upregulation of intestinal epithelial tight junction barrier is mediated by TLR-2/TLR-6 receptor complex activation of occludin gene

doi: 10.1038/s41522-025-00903-7

Figure Lengend Snippet: ( A ) Effect of BB1 on TLR-2/TLR-6 or TLR-2/TLR-1 heterodimer binding and signal-transduction pathway activation as determined by SEAP (secreted embryonic alkaline phosphatase) activity in TLR-1 or TLR-6 knock-out HEK-293 reporter cell lines (InvivoGen, San Diego, CA). BB1 produced a marked increase in SEAP activity in TLR-2/TLR-6 (TLR-1 -/- ), but not TLR-2/TLR-1 (TLR-6 -/- ), expressing HEK-293 cells. PAM3cBK4 (positive control) produced an increase in SEAP activity in TLR-2/TLR-1 (TLR-6 -/- ) HEK-293 cells. The siRNA knock-down of TLR-1 or TLR-6, but not TLR-2, inhibited the BB1-induced increase in occludin mRNA ( B ) and protein and densitometry analysis ( C, D ) and Caco-2 TEER ( E ) ∗∗∗∗ P < 0.0001 versus control, NS (not significant).

Article Snippet: Samples were incubated with anti-occludin antibody, anti-TOLLIP, or TLR-2 antibody (Cell Signaling Technology; Danvers, MA) overnight.

Techniques: Binding Assay, Transduction, Activation Assay, Activity Assay, Knock-Out, Produced, Expressing, Positive Control, Knockdown, Control

Journal: NPJ Biofilms and Microbiomes

Article Title: Probiotic bacteria Bifidobacterium bifidum upregulation of intestinal epithelial tight junction barrier is mediated by TLR-2/TLR-6 receptor complex activation of occludin gene

doi: 10.1038/s41522-025-00903-7

Figure Lengend Snippet: A BB1, but not BB4, treatment produced a focal aggregation of TLR-2 at the Caco-2 apical membrane surface and translocation of TOLLIP towards the apical membrane surface from the medial and basal regions of Caco-2 cells, as assessed by immunostaining. (TOLLIP, green; TLR-2 red; and DAPI blue). Scale bars = 10 μm. Quantification of TOLLIP apical translocation immunostaining intensity using ImageJ. ∗∗∗∗ P < 0.0001 versus control, NS (not significant) ( B ). The siRNA knockdown of TLR-2 or IRAK-1 inhibited the BB1-induced apical translocation of TOLLIP. The siRNA-induced of TOLLIP inhibited the BB1-induced increase in occludin mRNA ( C ) and protein expression ( D ), densitometry ( E ) and Caco-2 TEER ( F ). ∗∗∗∗ P < 0.0001 versus control, NS (not significant).

Article Snippet: Samples were incubated with anti-occludin antibody, anti-TOLLIP, or TLR-2 antibody (Cell Signaling Technology; Danvers, MA) overnight.

Techniques: Produced, Membrane, Translocation Assay, Immunostaining, Control, Knockdown, Expressing

Journal: NPJ Biofilms and Microbiomes

Article Title: Probiotic bacteria Bifidobacterium bifidum upregulation of intestinal epithelial tight junction barrier is mediated by TLR-2/TLR-6 receptor complex activation of occludin gene

doi: 10.1038/s41522-025-00903-7

Figure Lengend Snippet: A BB1, but not BB4, caused an increase in IRAK-1 phosphorylation (Thr209) as assessed by immunoblotting. The siRNA-induced knockdown of IRAK-1 inhibited the BB1-induced increase in occludin mRNA ( B ), occludin protein ( C ), densitometry ( D ) and Caco-2 TEER ( E ) (means ± SE, n = 4). ∗∗∗∗ P < 0.0001 versus control, NS (not significant). The siRNA-induced knockdown of TLR-2 or TLR-6, but not TLR-1, inhibited the BB1 increase in IRAK-1 phosphorylation, but had no effect on total IRAK-1 protein expression ( F ).

Article Snippet: Samples were incubated with anti-occludin antibody, anti-TOLLIP, or TLR-2 antibody (Cell Signaling Technology; Danvers, MA) overnight.

Techniques: Phospho-proteomics, Western Blot, Knockdown, Control, Expressing

Journal: NPJ Biofilms and Microbiomes

Article Title: Probiotic bacteria Bifidobacterium bifidum upregulation of intestinal epithelial tight junction barrier is mediated by TLR-2/TLR-6 receptor complex activation of occludin gene

doi: 10.1038/s41522-025-00903-7

Figure Lengend Snippet: A Oral -gastric gavage of BB1 [1 × 10 9 colony-forming units (CFUs)/mL], but not BB4, produced a decrease in mouse small intestinal permeability to dextran 10 kDa. P **** < 0.0001 versus control. BB1 administration caused an increase in occludin mRNA ( B ) and protein ( C ), densitometry ( D ) in the mouse intestinal tissue (24-hour treatment period). E BB1 administration also produced an increase in occludin expression and localization at the apical membrane surface and junctional areas, as assess by immunostaining. (Occludin, green; nuclei, blue). Scale bars = 10 μm ∗ P < 0.05, ∗∗∗ P < 0.001, and ∗∗∗∗ P < 0.0001, NS (not significant). ( E ). BB1 oral-gastric gavage administration produced a cytoplasmic-to-apical membrane translocation of TOLLIP in mouse intestinal epithelial cells ( F ). The BB1-induced apical membrane TOLLIP recruitment was inhibited in the TLR-2 ΔIEC mice (TOLLIP, green; occludin, red; (Leica SP8 confocal microscope).

Article Snippet: Samples were incubated with anti-occludin antibody, anti-TOLLIP, or TLR-2 antibody (Cell Signaling Technology; Danvers, MA) overnight.

Techniques: Produced, Permeability, Control, Expressing, Membrane, Immunostaining, Translocation Assay, Microscopy

Journal: NPJ Biofilms and Microbiomes

Article Title: Probiotic bacteria Bifidobacterium bifidum upregulation of intestinal epithelial tight junction barrier is mediated by TLR-2/TLR-6 receptor complex activation of occludin gene

doi: 10.1038/s41522-025-00903-7

Figure Lengend Snippet: A BB1 oral-gastric gavage produced an increase in small intestinal tissue expressions of TLR-2 and TLR-6 but not TLR-1. BB1 also produced an increase in apical membrane expression and localization of TLR-2 in mouse intestinal epithelial cells in the wild-type (WT) mice but not in the TLR-2 ΔIEC mice, as assessed by immunostaining (TLR-2, green; occludin, red; nuclei, blue; magnification 63x) ( B ). The BB1-induced increase in mouse intestinal tissue occludin mRNA ( C ) and protein ( D ), densitometry ( E ) and apical membrane and junctional localization of occludin (occludin, green; nuclei, blue) ( F ), and decrease in intestinal permeability to dextran 10 kDa ( G ) were inhibited in the TLR-2 ΔIEC mice. ∗∗∗∗ P < 0.0001 versus control, NS (not significant).

Article Snippet: Samples were incubated with anti-occludin antibody, anti-TOLLIP, or TLR-2 antibody (Cell Signaling Technology; Danvers, MA) overnight.

Techniques: Produced, Membrane, Expressing, Immunostaining, Permeability, Control

Journal: NPJ Biofilms and Microbiomes

Article Title: Probiotic bacteria Bifidobacterium bifidum upregulation of intestinal epithelial tight junction barrier is mediated by TLR-2/TLR-6 receptor complex activation of occludin gene

doi: 10.1038/s41522-025-00903-7

Figure Lengend Snippet: BB1 attaches to the apical membrane surface of the host intestinal epithelial cells by binding to the TLR-2/TLR-6 heterodimer complex and induces the phosphorylation of IRAK-1 and apical translocation of TOLLIP, which, in turn, leads to the down-steam activation of occludin gene and occludin protein synthesis and occludin-dependent upregulation of the intestinal TJ barrier.

Article Snippet: Samples were incubated with anti-occludin antibody, anti-TOLLIP, or TLR-2 antibody (Cell Signaling Technology; Danvers, MA) overnight.

Techniques: Membrane, Binding Assay, Phospho-proteomics, Translocation Assay, Serial Time-encoded Amplified Microscopy, Activation Assay

Review

tlr 2 antibody (Cell Signaling Technology Inc)

Structured Review

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