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anti pad2  (Proteintech)


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    Proteintech anti pad2
    Anti Pad2, supplied by Proteintech, used in various techniques. Bioz Stars score: 94/100, based on 70 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/anti pad2/product/Proteintech
    Average 94 stars, based on 70 article reviews
    anti pad2 - by Bioz Stars, 2026-05
    94/100 stars

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    anti pad2  (Proteintech)
    94
    Proteintech anti pad2
    Anti Pad2, supplied by Proteintech, 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/result/anti pad2/product/Proteintech
    Average 94 stars, based on 1 article reviews
    anti pad2 - by Bioz Stars, 2026-05
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    pad2 antibody  (Proteintech)
    94
    Proteintech pad2 antibody
    Generation of PAD2KO mice using the CRISPR-Cas9 systems and pathogenesis of TNBS-induced colitis in wild-type and PAD2KO mice. ( a ) The sequences produced for the generation of PAD2KO mice using the CRISPR-Cas9 systems, deficient in 61-bp on exon2 of the genomic <t>PAD2</t> DNA. ( b ) PAD2 mRNA level was determined in intraperitoneal macrophages by qRT-PCR normalized to TBP. Data were presented as the means ± S.E.M. (n = 8). ****P < 0.0001 vs. wild-type (WT) mice. ( c ) PAD2 expression was determined in intraperitoneal macrophages by western blotting normalized to β-actin expression. Data are presented as the means ± S.E.M. (n = 3). ***P < 0.001 vs. WT mice. ( d ) Body weight was measured daily in untreated-, vehicle (30% ethanol)-, or TNBS-injected WT and PAD2KO mice. On day 3, colon length was measured ( e ), representative images were obtained ( f ), and macroscopic analyses of the injured area were carried out using Image J ( g ). Scale bars are 10 mm. Histological changes were observed following H&E staining ( h ). ( i ) MPO activity was determined in those mice. Data are presented as the means ± S.E.M. (n = 8–9). #### P < 0.0001 vs. vehicle-injected mice (WT EtOH), and ****P < 0.0001, ***P < 0.001, *P < 0.05 vs. TNBS-injected mice (WT TNBS).
    Pad2 Antibody, supplied by Proteintech, 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/result/pad2 antibody/product/Proteintech
    Average 94 stars, based on 1 article reviews
    pad2 antibody - by Bioz Stars, 2026-05
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    pad2 antibodies  (Proteintech)
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    Proteintech pad2 antibodies
    Generation of PAD2KO mice using the CRISPR-Cas9 systems and pathogenesis of TNBS-induced colitis in wild-type and PAD2KO mice. ( a ) The sequences produced for the generation of PAD2KO mice using the CRISPR-Cas9 systems, deficient in 61-bp on exon2 of the genomic <t>PAD2</t> DNA. ( b ) PAD2 mRNA level was determined in intraperitoneal macrophages by qRT-PCR normalized to TBP. Data were presented as the means ± S.E.M. (n = 8). ****P < 0.0001 vs. wild-type (WT) mice. ( c ) PAD2 expression was determined in intraperitoneal macrophages by western blotting normalized to β-actin expression. Data are presented as the means ± S.E.M. (n = 3). ***P < 0.001 vs. WT mice. ( d ) Body weight was measured daily in untreated-, vehicle (30% ethanol)-, or TNBS-injected WT and PAD2KO mice. On day 3, colon length was measured ( e ), representative images were obtained ( f ), and macroscopic analyses of the injured area were carried out using Image J ( g ). Scale bars are 10 mm. Histological changes were observed following H&E staining ( h ). ( i ) MPO activity was determined in those mice. Data are presented as the means ± S.E.M. (n = 8–9). #### P < 0.0001 vs. vehicle-injected mice (WT EtOH), and ****P < 0.0001, ***P < 0.001, *P < 0.05 vs. TNBS-injected mice (WT TNBS).
    Pad2 Antibodies, supplied by Proteintech, 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/result/pad2 antibodies/product/Proteintech
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    rabbit anti pad2 mab  (Proteintech)
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    Proteintech rabbit anti pad2 mab
    Generation of PAD2KO mice using the CRISPR-Cas9 systems and pathogenesis of TNBS-induced colitis in wild-type and PAD2KO mice. ( a ) The sequences produced for the generation of PAD2KO mice using the CRISPR-Cas9 systems, deficient in 61-bp on exon2 of the genomic <t>PAD2</t> DNA. ( b ) PAD2 mRNA level was determined in intraperitoneal macrophages by qRT-PCR normalized to TBP. Data were presented as the means ± S.E.M. (n = 8). ****P < 0.0001 vs. wild-type (WT) mice. ( c ) PAD2 expression was determined in intraperitoneal macrophages by western blotting normalized to β-actin expression. Data are presented as the means ± S.E.M. (n = 3). ***P < 0.001 vs. WT mice. ( d ) Body weight was measured daily in untreated-, vehicle (30% ethanol)-, or TNBS-injected WT and PAD2KO mice. On day 3, colon length was measured ( e ), representative images were obtained ( f ), and macroscopic analyses of the injured area were carried out using Image J ( g ). Scale bars are 10 mm. Histological changes were observed following H&E staining ( h ). ( i ) MPO activity was determined in those mice. Data are presented as the means ± S.E.M. (n = 8–9). #### P < 0.0001 vs. vehicle-injected mice (WT EtOH), and ****P < 0.0001, ***P < 0.001, *P < 0.05 vs. TNBS-injected mice (WT TNBS).
    Rabbit Anti Pad2 Mab, supplied by Proteintech, used in various techniques. Bioz Stars score: 94/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    pad2  (Cell Signaling Technology Inc)
    93
    Cell Signaling Technology Inc pad2
    a Immunofluorescence analysis showing efficient uptake of Alexa Fluor 488-labeled CitH3 (5 µg/mL) by wild-type (WT) BMDMs but not by Tlr2⁻/⁻ BMDMs. Cellular intensity of Alexa-488 was quantified ( n = 5). Statistical analysis was performed using two-sided, Unpaired t test with Welch’s correction. b Cytokine profiling of pro- and anti-inflammatory mediators (IL-6, IL-1β, IL-10, TNFα, IFN-α, IFN-β) in supernatants of WT and Tlr2⁻/⁻ BMDMs treated with H3 or CitH3 peptides (15 µg/mL) for 16 h ( n = per group). LPS (200 ng/mL) served as positive control. Statistical analysis was performed using two-way ANOVA with Sidak’s multiple comparisons test. c Immunocytochemistry demonstrating nuclear translocation of <t>PAD2</t> in BMDMs after 1 h exposure to CitH3 peptide (15 µg/mL). LPS (200 ng/mL) served as a positive control. PAD2 was stained with Alexa Fluor 488 (green), and nuclei were counterstained with DAPI (blue). Quantification of PAD2 nuclear localization was analyzed by one-way ANOVA followed by Tukey’s multiple comparisons test ( n = 3 per group). d Subcellular fractionation analysis confirming significant nuclear localization and citrullination of PAD2 in WT BMDMs following treatment with H3, CitH3, or LPS. Quantification was analyzed using one-way ANOVA followed by Tukey’s multiple comparisons test ( n = per group). e In vitro citrullination assay showing MPB-tagged PAD2-mediated citrullination of H3 in a Ca²⁺-dependent manner. EGTA-mediated Ca²⁺ chelation eliminated CitH3 production. Similar results from three independent replicates. f In vitro citrullination assay showing PAD2 citrullination. PAD2 auto-citrullination was confirmed as His-tagged recombinant PAD2 co-incubated with MBP-tagged PAD2 resulted in PAD2 citrullination. PAD4 also promotes PAD2 citrullination. Quantification was performed by Image J, citrullination signals were normalized to corresponding input signals and further normalized to the matching EGTA-treated group in each replicate experiments. Statistical analysis was performed using two-sided t-test.Data are presented as mean ± SD. Statistical significance in all panels was determined as follows: * p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001. Source data are provided as a Source Data file.
    Pad2, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 93/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/pad2/product/Cell Signaling Technology Inc
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    rabbit anti human pad2 antibody  (Proteintech)
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    Proteintech rabbit anti human pad2 antibody
    Effect of AMF30a on in vitro proliferation of human corneal endothelial cells (hCECs). AMF30a (5µM, <t>PAD2</t> inhibitor, Cayman, Ann Arbor, Michigan) was applied for 48 h. ( A ) hCECs were identified using immunofluorescence staining of ZO1 and CD166. ( B - C ) Cell area of hCECs were evaluated using cell morphological pictures. Scale bar = 100 μm. ( D and E ) Cell proliferation was measured using CCK-8 cell viability assay and BrdU incorporation assay. ( F ) Cytotoxicity was evaluated using LDH cytotoxicity assay (1.002 ± 0.047 vs. 0.925 ± 0.018). ( G - I ) Cell cycle analysis was assessed using PI staining. (J and K) Wound healing assay was performed using the measurement of scratched ares. Scale bar = 500 μm. ( L and N ) Intracellular oxidative stress levels were measured using DCF-DA assay. Scale bar = 50 μm. * p < 0.05, ** p < 0.01 and **** p < 0.0001.
    Rabbit Anti Human Pad2 Antibody, supplied by Proteintech, 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/result/rabbit anti human pad2 antibody/product/Proteintech
    Average 94 stars, based on 1 article reviews
    rabbit anti human pad2 antibody - by Bioz Stars, 2026-05
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    rabbit anti pad2 antibody  (Santa Cruz Biotechnology)
    90
    Santa Cruz Biotechnology rabbit anti pad2 antibody
    Effect of AMF30a on in vitro proliferation of human corneal endothelial cells (hCECs). AMF30a (5µM, <t>PAD2</t> inhibitor, Cayman, Ann Arbor, Michigan) was applied for 48 h. ( A ) hCECs were identified using immunofluorescence staining of ZO1 and CD166. ( B - C ) Cell area of hCECs were evaluated using cell morphological pictures. Scale bar = 100 μm. ( D and E ) Cell proliferation was measured using CCK-8 cell viability assay and BrdU incorporation assay. ( F ) Cytotoxicity was evaluated using LDH cytotoxicity assay (1.002 ± 0.047 vs. 0.925 ± 0.018). ( G - I ) Cell cycle analysis was assessed using PI staining. (J and K) Wound healing assay was performed using the measurement of scratched ares. Scale bar = 500 μm. ( L and N ) Intracellular oxidative stress levels were measured using DCF-DA assay. Scale bar = 50 μm. * p < 0.05, ** p < 0.01 and **** p < 0.0001.
    Rabbit Anti Pad2 Antibody, supplied by Santa Cruz Biotechnology, used in various techniques. Bioz Stars score: 90/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/rabbit anti pad2 antibody/product/Santa Cruz Biotechnology
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    Image Search Results


    Generation of PAD2KO mice using the CRISPR-Cas9 systems and pathogenesis of TNBS-induced colitis in wild-type and PAD2KO mice. ( a ) The sequences produced for the generation of PAD2KO mice using the CRISPR-Cas9 systems, deficient in 61-bp on exon2 of the genomic PAD2 DNA. ( b ) PAD2 mRNA level was determined in intraperitoneal macrophages by qRT-PCR normalized to TBP. Data were presented as the means ± S.E.M. (n = 8). ****P < 0.0001 vs. wild-type (WT) mice. ( c ) PAD2 expression was determined in intraperitoneal macrophages by western blotting normalized to β-actin expression. Data are presented as the means ± S.E.M. (n = 3). ***P < 0.001 vs. WT mice. ( d ) Body weight was measured daily in untreated-, vehicle (30% ethanol)-, or TNBS-injected WT and PAD2KO mice. On day 3, colon length was measured ( e ), representative images were obtained ( f ), and macroscopic analyses of the injured area were carried out using Image J ( g ). Scale bars are 10 mm. Histological changes were observed following H&E staining ( h ). ( i ) MPO activity was determined in those mice. Data are presented as the means ± S.E.M. (n = 8–9). #### P < 0.0001 vs. vehicle-injected mice (WT EtOH), and ****P < 0.0001, ***P < 0.001, *P < 0.05 vs. TNBS-injected mice (WT TNBS).

    Journal: Scientific Reports

    Article Title: Peptidylarginine deiminase 2 contributes to pathogenesis in trinitrobenzenesulfonic acid-induced colitis through macrophage extracellular trap-independent pathways

    doi: 10.1038/s41598-025-24221-2

    Figure Lengend Snippet: Generation of PAD2KO mice using the CRISPR-Cas9 systems and pathogenesis of TNBS-induced colitis in wild-type and PAD2KO mice. ( a ) The sequences produced for the generation of PAD2KO mice using the CRISPR-Cas9 systems, deficient in 61-bp on exon2 of the genomic PAD2 DNA. ( b ) PAD2 mRNA level was determined in intraperitoneal macrophages by qRT-PCR normalized to TBP. Data were presented as the means ± S.E.M. (n = 8). ****P < 0.0001 vs. wild-type (WT) mice. ( c ) PAD2 expression was determined in intraperitoneal macrophages by western blotting normalized to β-actin expression. Data are presented as the means ± S.E.M. (n = 3). ***P < 0.001 vs. WT mice. ( d ) Body weight was measured daily in untreated-, vehicle (30% ethanol)-, or TNBS-injected WT and PAD2KO mice. On day 3, colon length was measured ( e ), representative images were obtained ( f ), and macroscopic analyses of the injured area were carried out using Image J ( g ). Scale bars are 10 mm. Histological changes were observed following H&E staining ( h ). ( i ) MPO activity was determined in those mice. Data are presented as the means ± S.E.M. (n = 8–9). #### P < 0.0001 vs. vehicle-injected mice (WT EtOH), and ****P < 0.0001, ***P < 0.001, *P < 0.05 vs. TNBS-injected mice (WT TNBS).

    Article Snippet: The membranes were then incubated with PAD2 antibody (1:500) (Proteintech, 12110-1-AP) and β-actin antibody (1:2000) (CST, #4970).

    Techniques: CRISPR, Produced, Quantitative RT-PCR, Expressing, Western Blot, Injection, Staining, Activity Assay

    Expression of inflammatory cytokines, chemokines, and PAD2 in PAD2KO mice. ( a ) TNFα, ( b ) IL-1β, ( c ) IL-12, ( d ) IL-23, ( e ) IFNγ, ( f ) CXCL2, and ( g ) PAD2 mRNA expression were determined using qRT-PCR in colon tissues of vehicle (30% ethanol)-, or TNBS-injected WT and PAD2KO mice on day 3, and mRNA levels were normalized to TBP. Data are presented as the means ± S.E.M. (n = 8–10). #### P < 0.0001, ### P < 0.001, ## P < 0.01 vs. vehicle-injected mice (WT EtOH), and ****P < 0.0001, *P < 0.05 vs. TNBS-injected mice (WT TNBS).

    Journal: Scientific Reports

    Article Title: Peptidylarginine deiminase 2 contributes to pathogenesis in trinitrobenzenesulfonic acid-induced colitis through macrophage extracellular trap-independent pathways

    doi: 10.1038/s41598-025-24221-2

    Figure Lengend Snippet: Expression of inflammatory cytokines, chemokines, and PAD2 in PAD2KO mice. ( a ) TNFα, ( b ) IL-1β, ( c ) IL-12, ( d ) IL-23, ( e ) IFNγ, ( f ) CXCL2, and ( g ) PAD2 mRNA expression were determined using qRT-PCR in colon tissues of vehicle (30% ethanol)-, or TNBS-injected WT and PAD2KO mice on day 3, and mRNA levels were normalized to TBP. Data are presented as the means ± S.E.M. (n = 8–10). #### P < 0.0001, ### P < 0.001, ## P < 0.01 vs. vehicle-injected mice (WT EtOH), and ****P < 0.0001, *P < 0.05 vs. TNBS-injected mice (WT TNBS).

    Article Snippet: The membranes were then incubated with PAD2 antibody (1:500) (Proteintech, 12110-1-AP) and β-actin antibody (1:2000) (CST, #4970).

    Techniques: Expressing, Quantitative RT-PCR, Injection

    a Immunofluorescence analysis showing efficient uptake of Alexa Fluor 488-labeled CitH3 (5 µg/mL) by wild-type (WT) BMDMs but not by Tlr2⁻/⁻ BMDMs. Cellular intensity of Alexa-488 was quantified ( n = 5). Statistical analysis was performed using two-sided, Unpaired t test with Welch’s correction. b Cytokine profiling of pro- and anti-inflammatory mediators (IL-6, IL-1β, IL-10, TNFα, IFN-α, IFN-β) in supernatants of WT and Tlr2⁻/⁻ BMDMs treated with H3 or CitH3 peptides (15 µg/mL) for 16 h ( n = per group). LPS (200 ng/mL) served as positive control. Statistical analysis was performed using two-way ANOVA with Sidak’s multiple comparisons test. c Immunocytochemistry demonstrating nuclear translocation of PAD2 in BMDMs after 1 h exposure to CitH3 peptide (15 µg/mL). LPS (200 ng/mL) served as a positive control. PAD2 was stained with Alexa Fluor 488 (green), and nuclei were counterstained with DAPI (blue). Quantification of PAD2 nuclear localization was analyzed by one-way ANOVA followed by Tukey’s multiple comparisons test ( n = 3 per group). d Subcellular fractionation analysis confirming significant nuclear localization and citrullination of PAD2 in WT BMDMs following treatment with H3, CitH3, or LPS. Quantification was analyzed using one-way ANOVA followed by Tukey’s multiple comparisons test ( n = per group). e In vitro citrullination assay showing MPB-tagged PAD2-mediated citrullination of H3 in a Ca²⁺-dependent manner. EGTA-mediated Ca²⁺ chelation eliminated CitH3 production. Similar results from three independent replicates. f In vitro citrullination assay showing PAD2 citrullination. PAD2 auto-citrullination was confirmed as His-tagged recombinant PAD2 co-incubated with MBP-tagged PAD2 resulted in PAD2 citrullination. PAD4 also promotes PAD2 citrullination. Quantification was performed by Image J, citrullination signals were normalized to corresponding input signals and further normalized to the matching EGTA-treated group in each replicate experiments. Statistical analysis was performed using two-sided t-test.Data are presented as mean ± SD. Statistical significance in all panels was determined as follows: * p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001. Source data are provided as a Source Data file.

    Journal: Nature Communications

    Article Title: A citrullinated histone H3 monoclonal antibody for immune modulation in sepsis

    doi: 10.1038/s41467-025-62788-6

    Figure Lengend Snippet: a Immunofluorescence analysis showing efficient uptake of Alexa Fluor 488-labeled CitH3 (5 µg/mL) by wild-type (WT) BMDMs but not by Tlr2⁻/⁻ BMDMs. Cellular intensity of Alexa-488 was quantified ( n = 5). Statistical analysis was performed using two-sided, Unpaired t test with Welch’s correction. b Cytokine profiling of pro- and anti-inflammatory mediators (IL-6, IL-1β, IL-10, TNFα, IFN-α, IFN-β) in supernatants of WT and Tlr2⁻/⁻ BMDMs treated with H3 or CitH3 peptides (15 µg/mL) for 16 h ( n = per group). LPS (200 ng/mL) served as positive control. Statistical analysis was performed using two-way ANOVA with Sidak’s multiple comparisons test. c Immunocytochemistry demonstrating nuclear translocation of PAD2 in BMDMs after 1 h exposure to CitH3 peptide (15 µg/mL). LPS (200 ng/mL) served as a positive control. PAD2 was stained with Alexa Fluor 488 (green), and nuclei were counterstained with DAPI (blue). Quantification of PAD2 nuclear localization was analyzed by one-way ANOVA followed by Tukey’s multiple comparisons test ( n = 3 per group). d Subcellular fractionation analysis confirming significant nuclear localization and citrullination of PAD2 in WT BMDMs following treatment with H3, CitH3, or LPS. Quantification was analyzed using one-way ANOVA followed by Tukey’s multiple comparisons test ( n = per group). e In vitro citrullination assay showing MPB-tagged PAD2-mediated citrullination of H3 in a Ca²⁺-dependent manner. EGTA-mediated Ca²⁺ chelation eliminated CitH3 production. Similar results from three independent replicates. f In vitro citrullination assay showing PAD2 citrullination. PAD2 auto-citrullination was confirmed as His-tagged recombinant PAD2 co-incubated with MBP-tagged PAD2 resulted in PAD2 citrullination. PAD4 also promotes PAD2 citrullination. Quantification was performed by Image J, citrullination signals were normalized to corresponding input signals and further normalized to the matching EGTA-treated group in each replicate experiments. Statistical analysis was performed using two-sided t-test.Data are presented as mean ± SD. Statistical significance in all panels was determined as follows: * p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001. Source data are provided as a Source Data file.

    Article Snippet: The primary antibodies used in immunocytochemistry assays: PAD2 (12110-1-AP), PAD4 (17373-1-AP), Rab5 (CST, 3547), EEA1 (CST, 3288), MPO (Abcam ab208670)

    Techniques: Immunofluorescence, Labeling, Positive Control, Immunocytochemistry, Translocation Assay, Staining, Fractionation, In Vitro, Recombinant, Incubation

    a LDH release from supernatants of BMDMs treated with increasing doses of CitH3 or unmodified H3 peptides for 24 h at 37 °C, showing CitH3-induced cytotoxicity ( n = per group). b Cell viability, assessed using the CCK8 assay, demonstrated significant reduction following CitH3 peptide treatment compared to the H3 peptide ( n = 5 per group). c BMDMs treated with 50 µg/mL CitH3 peptide and increasing doses of hCitH3-mAb or human IgG. LDH release was measured to evaluate cytotoxicity ( n = per group). d , Viability of BMDMs treated as in ( c ) was measured using the CCK8 assay. hCitH3-mAb significantly preserved cell viability compared to control IgG ( n = per group). e Immunoblot analysis of CitH3 in BMDMs treated with 15 μg/mL CitH3 or H3 peptides. Cells were washed three times prior to collection. And given that the CitH3 peptide is only ~30 amino acids in length, the bands observed at ~17 kDa are interpreted as endogenous cellular CitH3 protein detected at various time points. Similar results from three independent replicates. f Western blot analysis of citrullinated proteins in BMDMs from WT, Pad2⁻/⁻, and Pad2/4⁻/⁻ mice following 1 h treatment with 15 μg/mL CitH3 peptide. CitH3-induced citrullination was PAD2-dependent. Similar results from four independent replicates. g Quantification of IL-6, IL-1β, TNFα, IFN-α, and IFN-β levels in the supernatants of BMDMs treated with 25 μg/mL CitH3 peptide for 24 h ( n = 6 per group). hCitH3-mAb effectively reduced CitH3-induced cytokine elevation in a dose-dependent manner, whereas control IgG had no effect. Data are presented as mean ± SD. Statistical analysis: One-way ANOVA followed by Dunnett’s multiple comparisons test was applied throughout. Comparisons were made to the ‘0’ group in ( a –d ), and the CitH3-treated group in ( g ), as indicated. Significance thresholds: * p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001.Source data are provided as a Source Data file.

    Journal: Nature Communications

    Article Title: A citrullinated histone H3 monoclonal antibody for immune modulation in sepsis

    doi: 10.1038/s41467-025-62788-6

    Figure Lengend Snippet: a LDH release from supernatants of BMDMs treated with increasing doses of CitH3 or unmodified H3 peptides for 24 h at 37 °C, showing CitH3-induced cytotoxicity ( n = per group). b Cell viability, assessed using the CCK8 assay, demonstrated significant reduction following CitH3 peptide treatment compared to the H3 peptide ( n = 5 per group). c BMDMs treated with 50 µg/mL CitH3 peptide and increasing doses of hCitH3-mAb or human IgG. LDH release was measured to evaluate cytotoxicity ( n = per group). d , Viability of BMDMs treated as in ( c ) was measured using the CCK8 assay. hCitH3-mAb significantly preserved cell viability compared to control IgG ( n = per group). e Immunoblot analysis of CitH3 in BMDMs treated with 15 μg/mL CitH3 or H3 peptides. Cells were washed three times prior to collection. And given that the CitH3 peptide is only ~30 amino acids in length, the bands observed at ~17 kDa are interpreted as endogenous cellular CitH3 protein detected at various time points. Similar results from three independent replicates. f Western blot analysis of citrullinated proteins in BMDMs from WT, Pad2⁻/⁻, and Pad2/4⁻/⁻ mice following 1 h treatment with 15 μg/mL CitH3 peptide. CitH3-induced citrullination was PAD2-dependent. Similar results from four independent replicates. g Quantification of IL-6, IL-1β, TNFα, IFN-α, and IFN-β levels in the supernatants of BMDMs treated with 25 μg/mL CitH3 peptide for 24 h ( n = 6 per group). hCitH3-mAb effectively reduced CitH3-induced cytokine elevation in a dose-dependent manner, whereas control IgG had no effect. Data are presented as mean ± SD. Statistical analysis: One-way ANOVA followed by Dunnett’s multiple comparisons test was applied throughout. Comparisons were made to the ‘0’ group in ( a –d ), and the CitH3-treated group in ( g ), as indicated. Significance thresholds: * p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001.Source data are provided as a Source Data file.

    Article Snippet: The primary antibodies used in immunocytochemistry assays: PAD2 (12110-1-AP), PAD4 (17373-1-AP), Rab5 (CST, 3547), EEA1 (CST, 3288), MPO (Abcam ab208670)

    Techniques: CCK-8 Assay, Control, Western Blot

    Effect of AMF30a on in vitro proliferation of human corneal endothelial cells (hCECs). AMF30a (5µM, PAD2 inhibitor, Cayman, Ann Arbor, Michigan) was applied for 48 h. ( A ) hCECs were identified using immunofluorescence staining of ZO1 and CD166. ( B - C ) Cell area of hCECs were evaluated using cell morphological pictures. Scale bar = 100 μm. ( D and E ) Cell proliferation was measured using CCK-8 cell viability assay and BrdU incorporation assay. ( F ) Cytotoxicity was evaluated using LDH cytotoxicity assay (1.002 ± 0.047 vs. 0.925 ± 0.018). ( G - I ) Cell cycle analysis was assessed using PI staining. (J and K) Wound healing assay was performed using the measurement of scratched ares. Scale bar = 500 μm. ( L and N ) Intracellular oxidative stress levels were measured using DCF-DA assay. Scale bar = 50 μm. * p < 0.05, ** p < 0.01 and **** p < 0.0001.

    Journal: Scientific Reports

    Article Title: AMF30a promotes survival and function of human corneal endothelial cells by regulating TGF-β/ROCK/HIPPO pathway

    doi: 10.1038/s41598-025-13656-2

    Figure Lengend Snippet: Effect of AMF30a on in vitro proliferation of human corneal endothelial cells (hCECs). AMF30a (5µM, PAD2 inhibitor, Cayman, Ann Arbor, Michigan) was applied for 48 h. ( A ) hCECs were identified using immunofluorescence staining of ZO1 and CD166. ( B - C ) Cell area of hCECs were evaluated using cell morphological pictures. Scale bar = 100 μm. ( D and E ) Cell proliferation was measured using CCK-8 cell viability assay and BrdU incorporation assay. ( F ) Cytotoxicity was evaluated using LDH cytotoxicity assay (1.002 ± 0.047 vs. 0.925 ± 0.018). ( G - I ) Cell cycle analysis was assessed using PI staining. (J and K) Wound healing assay was performed using the measurement of scratched ares. Scale bar = 500 μm. ( L and N ) Intracellular oxidative stress levels were measured using DCF-DA assay. Scale bar = 50 μm. * p < 0.05, ** p < 0.01 and **** p < 0.0001.

    Article Snippet: Primary antibodies were as follows: rabbit anti-human CD166 antibody (ab109215, Abcam, 1∶500 dilution), rabbit anti-human ZO-1 antibody (sc-10804, Santa Cruz, 1∶500 dilution), mouse anti-human ERK1/2 antibody (sc-514032, Santa Cruz, 1∶500 dilution); mouse anti-human pERK1/2 antibody (sc-136521, Santa Cruz, 1∶500 dilution); mouse anti-human YAP antibody (sc-376830, 1∶500 dilution); rabbit anti-pYAP antibody (PA5-17481, Invitrogen, 1∶500 dilution); rabbit anti-human PAD2 antibody (12110-1-AP, Proteintech, 1∶500 dilution); mouse anti-ATF4 antibody (sc-390063, 1∶500 dilution); mouse anti-human ROCK1 antibody (sc-17794, 1∶500 dilution); mouse anti-ROCK2 antibody (sc-398519, 1∶500 dilution); or rabbit anti-GAPDH antibody (LF-PA0212, Abfrontier, 1∶5000 dilution).

    Techniques: In Vitro, Immunofluorescence, Staining, CCK-8 Assay, Viability Assay, BrdU Incorporation Assay, LDH Cytotoxicity Assay, Cell Cycle Assay, Wound Healing Assay

    Effect of cytokine on human corneal endothelial cells. ( A ) Immunofluorescence staining of PAD2 shows the distribution of PAD2. Scale bar = 100 μm. ( B and C ) Western blot of PAD2 was used to evaluate the PAD2 levels. ( D and E ) ATF4 levels were evaluated using western blot. ( F ) Immunofluorescence staining of peptidyl-citrulline shows the intensity and distribution of peptidyl-citrulline. Scale bar = 100 μm. * p < 0.05.

    Journal: Scientific Reports

    Article Title: AMF30a promotes survival and function of human corneal endothelial cells by regulating TGF-β/ROCK/HIPPO pathway

    doi: 10.1038/s41598-025-13656-2

    Figure Lengend Snippet: Effect of cytokine on human corneal endothelial cells. ( A ) Immunofluorescence staining of PAD2 shows the distribution of PAD2. Scale bar = 100 μm. ( B and C ) Western blot of PAD2 was used to evaluate the PAD2 levels. ( D and E ) ATF4 levels were evaluated using western blot. ( F ) Immunofluorescence staining of peptidyl-citrulline shows the intensity and distribution of peptidyl-citrulline. Scale bar = 100 μm. * p < 0.05.

    Article Snippet: Primary antibodies were as follows: rabbit anti-human CD166 antibody (ab109215, Abcam, 1∶500 dilution), rabbit anti-human ZO-1 antibody (sc-10804, Santa Cruz, 1∶500 dilution), mouse anti-human ERK1/2 antibody (sc-514032, Santa Cruz, 1∶500 dilution); mouse anti-human pERK1/2 antibody (sc-136521, Santa Cruz, 1∶500 dilution); mouse anti-human YAP antibody (sc-376830, 1∶500 dilution); rabbit anti-pYAP antibody (PA5-17481, Invitrogen, 1∶500 dilution); rabbit anti-human PAD2 antibody (12110-1-AP, Proteintech, 1∶500 dilution); mouse anti-ATF4 antibody (sc-390063, 1∶500 dilution); mouse anti-human ROCK1 antibody (sc-17794, 1∶500 dilution); mouse anti-ROCK2 antibody (sc-398519, 1∶500 dilution); or rabbit anti-GAPDH antibody (LF-PA0212, Abfrontier, 1∶5000 dilution).

    Techniques: Immunofluorescence, Staining, Western Blot

    Schematic diagram summarizing PAD2-mediated signaling in hCECs.

    Journal: Scientific Reports

    Article Title: AMF30a promotes survival and function of human corneal endothelial cells by regulating TGF-β/ROCK/HIPPO pathway

    doi: 10.1038/s41598-025-13656-2

    Figure Lengend Snippet: Schematic diagram summarizing PAD2-mediated signaling in hCECs.

    Article Snippet: Primary antibodies were as follows: rabbit anti-human CD166 antibody (ab109215, Abcam, 1∶500 dilution), rabbit anti-human ZO-1 antibody (sc-10804, Santa Cruz, 1∶500 dilution), mouse anti-human ERK1/2 antibody (sc-514032, Santa Cruz, 1∶500 dilution); mouse anti-human pERK1/2 antibody (sc-136521, Santa Cruz, 1∶500 dilution); mouse anti-human YAP antibody (sc-376830, 1∶500 dilution); rabbit anti-pYAP antibody (PA5-17481, Invitrogen, 1∶500 dilution); rabbit anti-human PAD2 antibody (12110-1-AP, Proteintech, 1∶500 dilution); mouse anti-ATF4 antibody (sc-390063, 1∶500 dilution); mouse anti-human ROCK1 antibody (sc-17794, 1∶500 dilution); mouse anti-ROCK2 antibody (sc-398519, 1∶500 dilution); or rabbit anti-GAPDH antibody (LF-PA0212, Abfrontier, 1∶5000 dilution).

    Techniques:

    Effect of AMF30a on in vitro proliferation of human corneal endothelial cells (hCECs). AMF30a (5µM, PAD2 inhibitor, Cayman, Ann Arbor, Michigan) was applied for 48 h. ( A ) hCECs were identified using immunofluorescence staining of ZO1 and CD166. ( B - C ) Cell area of hCECs were evaluated using cell morphological pictures. Scale bar = 100 μm. ( D and E ) Cell proliferation was measured using CCK-8 cell viability assay and BrdU incorporation assay. ( F ) Cytotoxicity was evaluated using LDH cytotoxicity assay (1.002 ± 0.047 vs. 0.925 ± 0.018). ( G - I ) Cell cycle analysis was assessed using PI staining. (J and K) Wound healing assay was performed using the measurement of scratched ares. Scale bar = 500 μm. ( L and N ) Intracellular oxidative stress levels were measured using DCF-DA assay. Scale bar = 50 μm. * p < 0.05, ** p < 0.01 and **** p < 0.0001.

    Journal: Scientific Reports

    Article Title: AMF30a promotes survival and function of human corneal endothelial cells by regulating TGF-β/ROCK/HIPPO pathway

    doi: 10.1038/s41598-025-13656-2

    Figure Lengend Snippet: Effect of AMF30a on in vitro proliferation of human corneal endothelial cells (hCECs). AMF30a (5µM, PAD2 inhibitor, Cayman, Ann Arbor, Michigan) was applied for 48 h. ( A ) hCECs were identified using immunofluorescence staining of ZO1 and CD166. ( B - C ) Cell area of hCECs were evaluated using cell morphological pictures. Scale bar = 100 μm. ( D and E ) Cell proliferation was measured using CCK-8 cell viability assay and BrdU incorporation assay. ( F ) Cytotoxicity was evaluated using LDH cytotoxicity assay (1.002 ± 0.047 vs. 0.925 ± 0.018). ( G - I ) Cell cycle analysis was assessed using PI staining. (J and K) Wound healing assay was performed using the measurement of scratched ares. Scale bar = 500 μm. ( L and N ) Intracellular oxidative stress levels were measured using DCF-DA assay. Scale bar = 50 μm. * p < 0.05, ** p < 0.01 and **** p < 0.0001.

    Article Snippet: The cells were treated overnight with either mouse anti-E-cadherin antibody (sc-8426; Santa Cruz Biotechnology, Santa Cruz, CA, USA) or mouse anti-YAP antibody (sc-376830; Santa Cruz Biotechnology), rabbit anti-PAD2 antibody (sc-293271; Santa Cruz Biotechnology), mouse anti-peptidyl-citrulline antibody (MABN328, Merck Millipore, Feltham, UK), and rabbit anti-NF-κB antibody (ab7970, Abcam, 1:100) at 4 °C and then rinsed with PBS.

    Techniques: In Vitro, Immunofluorescence, Staining, CCK-8 Assay, Viability Assay, BrdU Incorporation Assay, LDH Cytotoxicity Assay, Cell Cycle Assay, Wound Healing Assay

    Effect of cytokine on human corneal endothelial cells. ( A ) Immunofluorescence staining of PAD2 shows the distribution of PAD2. Scale bar = 100 μm. ( B and C ) Western blot of PAD2 was used to evaluate the PAD2 levels. ( D and E ) ATF4 levels were evaluated using western blot. ( F ) Immunofluorescence staining of peptidyl-citrulline shows the intensity and distribution of peptidyl-citrulline. Scale bar = 100 μm. * p < 0.05.

    Journal: Scientific Reports

    Article Title: AMF30a promotes survival and function of human corneal endothelial cells by regulating TGF-β/ROCK/HIPPO pathway

    doi: 10.1038/s41598-025-13656-2

    Figure Lengend Snippet: Effect of cytokine on human corneal endothelial cells. ( A ) Immunofluorescence staining of PAD2 shows the distribution of PAD2. Scale bar = 100 μm. ( B and C ) Western blot of PAD2 was used to evaluate the PAD2 levels. ( D and E ) ATF4 levels were evaluated using western blot. ( F ) Immunofluorescence staining of peptidyl-citrulline shows the intensity and distribution of peptidyl-citrulline. Scale bar = 100 μm. * p < 0.05.

    Article Snippet: The cells were treated overnight with either mouse anti-E-cadherin antibody (sc-8426; Santa Cruz Biotechnology, Santa Cruz, CA, USA) or mouse anti-YAP antibody (sc-376830; Santa Cruz Biotechnology), rabbit anti-PAD2 antibody (sc-293271; Santa Cruz Biotechnology), mouse anti-peptidyl-citrulline antibody (MABN328, Merck Millipore, Feltham, UK), and rabbit anti-NF-κB antibody (ab7970, Abcam, 1:100) at 4 °C and then rinsed with PBS.

    Techniques: Immunofluorescence, Staining, Western Blot

    Schematic diagram summarizing PAD2-mediated signaling in hCECs.

    Journal: Scientific Reports

    Article Title: AMF30a promotes survival and function of human corneal endothelial cells by regulating TGF-β/ROCK/HIPPO pathway

    doi: 10.1038/s41598-025-13656-2

    Figure Lengend Snippet: Schematic diagram summarizing PAD2-mediated signaling in hCECs.

    Article Snippet: The cells were treated overnight with either mouse anti-E-cadherin antibody (sc-8426; Santa Cruz Biotechnology, Santa Cruz, CA, USA) or mouse anti-YAP antibody (sc-376830; Santa Cruz Biotechnology), rabbit anti-PAD2 antibody (sc-293271; Santa Cruz Biotechnology), mouse anti-peptidyl-citrulline antibody (MABN328, Merck Millipore, Feltham, UK), and rabbit anti-NF-κB antibody (ab7970, Abcam, 1:100) at 4 °C and then rinsed with PBS.

    Techniques: