rabbit pab against mcherry  (Proteintech)

Journal: bioRxiv

Article Title: ASFV early protein p30 suppresses antiviral type I IFN induction by targeting TRIM21 and RIG-I like receptor signaling adaptor MAVS

doi: 10.64898/2026.03.26.714469

Figure Lengend Snippet: Interactions exist between ASFV p30, TRIM21 and MAVS. ( A ) 293T cells were co-transfected with GFP-MAVS and Flag-TRIM21 for 24 h. Immunoprecipitation was performed using the rabbit anti-GFP mAb to confirm the interaction between exogenous MAVS and TRIM21. ( B ) 293T cells were co-transfected with mCherry-MAVS, Flag-TRIM21, and p30-GFP for 24 h. Immunoprecipitation was performed using rabbit mCherry pAb to confirm the interactions between exogenous MAVS, TRIM21 and p30. ( C ) MA104 cells were infected with ASFV for 72 h. Immunoprecipitation were performed using rabbit anti-MAVS mAb to confirm the interactions between endogenous MAVS, TRIM21 and p30. ( D ) The purified p30, TRIM21 and MAVS recombinant proteins were confirmed by SDS-PAGE and labeled with red boxes. ( E-G ) The interactions between purified proteins p30 and TRIM21, p30 and MAVS, and MAVS and TRIM21 verified through immunoprecipitations.

Article Snippet: The rabbit anti-TRIM21 mAb, mouse anti-TRIM21 mAb, rabbit anti-MAVS mAb, rabbit anti-IRF3 mAb, rabbit anti-Flag mAb, rabbit anti-HA mAb and rabbit mCherry pAb were all from ProteinTech (Wuhan, China).

Techniques: Transfection, Immunoprecipitation, Infection, Purification, Recombinant, SDS Page, Labeling

Journal: bioRxiv

Article Title: ASFV early protein p30 suppresses antiviral type I IFN induction by targeting TRIM21 and RIG-I like receptor signaling adaptor MAVS

doi: 10.64898/2026.03.26.714469

Figure Lengend Snippet: ASFV p30 inhibits TRIM21 upregulated MAVS signaling. ( A ) 293T cells were co-transfected mCherry-MAVS, Flag-TRIM21 and p30-GFP, as indicated, for 24 h. The downstream ISG56 and IFN-β mRNA levels activated by MAVS in the presence of TRIM21 were analyzed using RT-qPCR. ( B ) 3D4/21 cells were co-transfected with Flag-TRIM21 and p30-GFP, as indicated, for 12 h, and stimulated by transfection of poly (I:C) for 12 h. The downstream ISG56 and IFN-β mRNA levels activated by poly (I:C) in the presence of TRIM21 were analyzed using RT-qPCR. ( C ) 293T cells were co-transfected with mCherry-MAVS, Flag-TRIM21 and p30-GFP, plus ISRE Fluc and Rluc reporters, as indicated, for 24 h, and ISRE promoter activity was measured via dual luciferase reporter assay. ( D and E ) 3D4/21 cells were co-transfected with Flag-TRIM21 and p30-GFP for 12 h, as indicated. Cells were infected with VSV (D) or stimulated with transfection of poly (I:C) (E), followed by Western blotting for detection of cell signaling activation. ( F ) PAMs were transfected with p30 siRNA or control siRNA for 24 h, stimulated with transfection of poly (I:C) for 6 h, and then infected with ASFV for 48 h. The cell signaling and ASFV replication were detected by Western blotting.

Article Snippet: The rabbit anti-TRIM21 mAb, mouse anti-TRIM21 mAb, rabbit anti-MAVS mAb, rabbit anti-IRF3 mAb, rabbit anti-Flag mAb, rabbit anti-HA mAb and rabbit mCherry pAb were all from ProteinTech (Wuhan, China).

Techniques: Transfection, Quantitative RT-PCR, Activity Assay, Luciferase, Reporter Assay, Infection, Western Blot, Activation Assay, Control

Journal: bioRxiv

Article Title: ASFV early protein p30 suppresses antiviral type I IFN induction by targeting TRIM21 and RIG-I like receptor signaling adaptor MAVS

doi: 10.64898/2026.03.26.714469

Figure Lengend Snippet: ASFV p30 inhibits TRIM21 mediated ubiquitination of MAVS K27 ubiquitination. ( A ) 293T cells were co-transfected with Flag-TRIM21, mCherry-MAVS and HA-Ub for 24 h and then treated with MG-132 (10 μm) for 6 h. ( B ) 293T cells were co-transfected with Flag-TRIM21, mCherry-MAVS, p30-GFP, and HA-Ub for 24 h and then treated with MG-132 (10 μm) for 6 h. Cell samples were collected and immunoprecipitation were used for analysis of MAVS ubiquitination. ( C ) 293T cells were co-transfected with Flag-TRIM21, mCherry-MAVS and HA-Ub-K27 for 24 h and then treated with MG-132 (10 μm) for 6 h. ( D ) 293T cells were co-transfected with Flag-TRIM21, mCherry-MAVS, p30-myc, and HA-Ub-K27 for 24 h and then treated with MG-132 (10 μm) for 6 h. Cell samples were collected and immunoprecipitation were used for analysis of MAVS K27 ubiquitination.

Article Snippet: The rabbit anti-TRIM21 mAb, mouse anti-TRIM21 mAb, rabbit anti-MAVS mAb, rabbit anti-IRF3 mAb, rabbit anti-Flag mAb, rabbit anti-HA mAb and rabbit mCherry pAb were all from ProteinTech (Wuhan, China).

Techniques: Ubiquitin Proteomics, Transfection, Immunoprecipitation

Journal: bioRxiv

Article Title: DDX3X syndrome mutations lock DDX3X-RNA conformational states to drive persistent pathological condensation and neuronal death

doi: 10.64898/2026.01.29.702492

Figure Lengend Snippet: A . Schematic representation of DDX3X protein showing pathogenic mutations identified in DDX3X syndrome patients (bold- multiple occurrences, blue- polymicrogyria, brown- male patients). B . Schematic representation of DDX3X functional domains indicating critical DDX3X syndrome-associated mutations selected based on their predicted structural impact. Blue, RNA binding sites; Red, ATP binding and hydrolysis regions; Magenta, regions of interaction between ATP binding and RNA binding residues. C . Ribbon structural representation of DDX3X (dark grey) bound to dsRNA (red) (PDB ID: 6O5F), displaying surface accessibility and RNA interface proximity of DDX3X syndrome mutations. Mutated residues are color-coded in the represented structure. D . Heat map depicting the AlphaMissense prediction for the likely pathogenicity of selected DDX3X syndrome missense mutations, with dark red being the likely pathogenic and white being the likely benign mutation. E . Table depicting selected mutations in DDX3X that have been associated with different cancer types. F . Immunoblotting analysis of DDX3X showing the impact of DDX3X syndrome mutations on protein expression in Neuro2a cells. G . Fluorescence images of N2a cells expressing DDX3X-mCherry with DDX3X syndrome mutations, captured using Incucyte imager.

Article Snippet: Full length DDX3X-mCherry construct (pReceiver-M56-DDX3X-mCherry ) was procured from GeneCopoeia.

Techniques: Functional Assay, RNA Binding Assay, Binding Assay, Mutagenesis, Western Blot, Expressing, Fluorescence

Journal: bioRxiv

Article Title: DDX3X syndrome mutations lock DDX3X-RNA conformational states to drive persistent pathological condensation and neuronal death

doi: 10.64898/2026.01.29.702492

Figure Lengend Snippet: A . Immunoblot analysis showing levels of DDX3X protein in HeLa WT & Ddx3x -/- cells. B . Representative confocal microscopy images of HeLa cells expressing WT or mutant DDX3X-mCherry constructs and subjected to SA (200µM) stress and immunostained for stress granule marker G3BP1 (green) and DAPI (blue). Scale bar=10µm. C-E . Quantification of numbers of DDX3X (red), G3BP1 (green), and DDX3X-G3BP1 (yellow) granules per cell quantified from ≥50 cells across three independent experiments exhibiting DDX3X colocalising with stress granules in HeLa cells. ****p < 0.0001, ***p = 0.0008 (DDX3X granules/cell), **p = 0.0022 (WT vs I190S), **p = 0.0028 (WT vs T323I), **p = 0.0031 (WT vs L556S) (G3BP1 granules/cells), ****p < 0.0001, **p = 0.0034 (G3BP1-DDX3X colocalised granules/cells) (One-way ANOVA test). Data shown are mean ± SEM.

Article Snippet: Full length DDX3X-mCherry construct (pReceiver-M56-DDX3X-mCherry ) was procured from GeneCopoeia.

Techniques: Western Blot, Confocal Microscopy, Expressing, Mutagenesis, Construct, Marker

Journal: bioRxiv

Article Title: DDX3X syndrome mutations lock DDX3X-RNA conformational states to drive persistent pathological condensation and neuronal death

doi: 10.64898/2026.01.29.702492

Figure Lengend Snippet: A. Representative confocal images of N2a cells expressing WT or mutant DDX3X-mCherry subjected to SA (100µM) stress for 2 hours and immunostained for stress granule marker G3BP1 (green) and DAPI (blue). Scale bar=10µm. B-D . Quantification of numbers of DDX3X (red), G3BP1 (green) and DDX3X-G3BP1 (yellow) granules per cell from ≥ 30cells across three independent experiments exhibiting DDX3X colocalising with stress granules in N2a cells. ****p < 0.0001, ***p = 0.0003, *p = 0.0181 (DDX3X granules/cell), ****p < 0.0001, ***p = 0.0001 (WT VS T198P), ***p = 0.0003 (WT vs L559H),**p = 0.0016 (G3BP1 granules/cells), ****p < 0.0001, ***p = 0.0001, **p = 0.0013 (WT vs I190S), **p = 0.0024 (WT vs L559H) (G3BP1-DDX3X colocalised granules/cells) (One-way ANOVA test). Data shown are mean ± SEM.

Article Snippet: Full length DDX3X-mCherry construct (pReceiver-M56-DDX3X-mCherry ) was procured from GeneCopoeia.

Techniques: Expressing, Mutagenesis, Marker

Journal: bioRxiv

Article Title: DDX3X syndrome mutations lock DDX3X-RNA conformational states to drive persistent pathological condensation and neuronal death

doi: 10.64898/2026.01.29.702492

Figure Lengend Snippet: A . Schematic showing the protocol used for monitoring persistent granules in cells. Cells were treated with sodium arsenite (SA), followed by removing SA and supplementing fresh media (wash-off, recovery period) to monitor persistent granules. B . Representative confocal microscopy images of HeLa cells expressing WT or mutant DDX3X-mCherry showing stress granules (SGs) upon SA treatment and after wash-off (recovery), immunostained for G3BP1 (green) and DAPI (blue). Scale bar = 10µm. C-D . Quantification of the number of DDX3X (red) and DDX3X-G3BP1 (yellow) granules per cell from ≥ 30 cells across three independent experiments to monitor the formation of persistent DDX3X-SGs due to DDX3X syndrome mutations in HeLa cells. ****p < 0.0001, ***p = 0.0002, **p = 0.0073 (WT vs WT-WO), **p = 0.0046 (R475G vs R475G-WO), ns = not significant (DDX3X granules/cell), ****p < 0.0001, ***p = 0.0001, **p = 0.0096 (G3BP1-DDX3X granules/cell) (one-way ANOVA test). Data shown are mean ± SEM. E . Representative confocal microscopy images of N2a cells expressing WT or mutant DDX3X-mCherry showing SGs upon SA treatment and after wash-off (recovery), immunostained for G3BP1 (green) and DAPI (blue). Scale bar = 10µm. F-G . Quantification of the number of DDX3X (red) and DDX3X-G3BP1 (yellow) granules per cell from ≥ 50 cells across three independent experiments to monitor the formation of persistent DDX3X-SGs due to DDX3X syndrome mutations in N2a cells., ***p = 0.0006, **p = 0.0033 (WT vs WT-WO), **p = 0.0020 (F182V vs F182V-WO), ns = non-significant (DDX3X granules/cell), ****p < 0.0001, ***p = 0.0003 (G3BP1-DDX3X granules/cell) (One-way ANOVA test). Data shown are mean ± SEM.

Article Snippet: Full length DDX3X-mCherry construct (pReceiver-M56-DDX3X-mCherry ) was procured from GeneCopoeia.

Techniques: Confocal Microscopy, Expressing, Mutagenesis

Journal: bioRxiv

Article Title: DDX3X syndrome mutations lock DDX3X-RNA conformational states to drive persistent pathological condensation and neuronal death

doi: 10.64898/2026.01.29.702492

Figure Lengend Snippet: A . Real-time time-lapse confocal images of HeLa cells expressing WT or mutant DDX3X-mCherry and subjected to SA (200µM) stress for 2 hours. A single SG (indicated by white dashed circle, as the region of interest) was photobleached and fluorescence recovery was recorded over 180s using confocal microscopy. B . The relative fluorescence intensity of mCherry (tagged to DDX3X) in SGs in HeLa cells before and after photobleaching as a function of time. Curves are representative of four independent experiments. C. Real-time time-lapse confocal images of N2a cells expressing WT or mutant DDX3X-mCherry and subjected to SA (100µM) stress for 2 hours. A single SG (indicated by a dashed circle, as the ROI) was photobleached, and fluorescence recovery was recorded over 150s using confocal microscopy. D. The relative fluorescence intensity of mCherry (tagged to DDX3X) in SGs in N2a cells before and after photobleaching as a function of time. Curves are representative of five independent experiments.

Article Snippet: Full length DDX3X-mCherry construct (pReceiver-M56-DDX3X-mCherry ) was procured from GeneCopoeia.

Techniques: Expressing, Mutagenesis, Fluorescence, Confocal Microscopy

Journal: bioRxiv

Article Title: DDX3X syndrome mutations lock DDX3X-RNA conformational states to drive persistent pathological condensation and neuronal death

doi: 10.64898/2026.01.29.702492

Figure Lengend Snippet: A. Schematic of the assay protocol to assess the effect of persistent SGs on global translation and cell death. B. Representative polysome profiles (A260 absorbance) of DDX3X syndrome mutants expressing N2a cell extracts treated with SA (100uM) followed by wash-off and recovery in SA-free media, indicating sedimentation (through 10%-50% sucrose gradient) of 40S & 60S ribosomal subunits, 80S monosomes and polysomes (n=3). C. Quantification of DDX3X-dependent mRNAs in the polysome fractions plotted as relative log2-fold change ratio of translated/untranslated fraction using gene-specific primers normalised to GAPDH (internal control). ****p < 0.0001, ***p = 0.0007 (WT vs L559H, RPL36A gene), ***p = 0.0002 (WT vs L559H, RPL13 gene), **p = 0.0034 (WT vs I190S, EIF3I gene), **p = 0.0024 (WT vs L556S, STAT1 gene), **p = 0.0035 (WT vs I190S, TOPBP1 gene), **p = 0.0016 (WT vs L556S, TOPBP1), **p = 0.00365 (WT vs F182V, GNB2 gene), *p = 0.0347 (One-way ANOVA). Data shown are mean ± SEM. D. Real-time cell death analysis by Sytox green staining of N2a cells expressing WT or mutant DDX3X-mCherry constructs treated with SA **p = 0.0079, *p = 0.0432 (WT vs I190S), *p = 0.0306 (WT vs L559H) (two-way ANOVA test). Data shown are mean ± SEM. E. Real-time cell death analysis by Sytox green staining of N2a cells expressing WT or mutant DDX3X constructs treated with Aβ peptide with or without SA treatment and wash off. ****p < 0.0001, ***p = 0.0004, **p = 0.0068 (two-way ANOVA test). Data shown are mean ± SEM. F. Cell death measurement by Sytox green staining of N2a cells expressing WT or mutant DDX3X treated with SA, followed by TNF and zVAD treatment. ****p < 0.0001, **p = 0.0020 (WT vs T198P), **p = 0.0038 (WT vs R480G), **p = 0.0053 (WT vs L556S), *p = 0.0182 (two-way ANOVA test). Data shown are mean ± SEM.

Article Snippet: Full length DDX3X-mCherry construct (pReceiver-M56-DDX3X-mCherry ) was procured from GeneCopoeia.

Techniques: Expressing, Sedimentation, Control, Staining, Mutagenesis, Construct

Journal: bioRxiv

Article Title: DDX3X syndrome mutations lock DDX3X-RNA conformational states to drive persistent pathological condensation and neuronal death

doi: 10.64898/2026.01.29.702492

Figure Lengend Snippet: A. Cell death measurement by Sytox green staining of N2a cells ectopically expressing WT or mutant DDX3X-mCherry. ns = non-significant (Two-way ANOVA test). Data shown are mean ± SEM. B. Cell death measurement by Sytox green staining of N2a cells ectopically expressing WT or mutant DDX3X-mCherry treated with TNF and zVAD. Ns = non-significant, (Two-way ANOVA test). Data shown are mean ± SEM.

Article Snippet: Full length DDX3X-mCherry construct (pReceiver-M56-DDX3X-mCherry ) was procured from GeneCopoeia.

Techniques: Staining, Expressing, Mutagenesis

Journal: bioRxiv

Article Title: DDX3X syndrome mutations lock DDX3X-RNA conformational states to drive persistent pathological condensation and neuronal death

doi: 10.64898/2026.01.29.702492

Figure Lengend Snippet: A . Schematic showing the protocol used for the cross-seeding kinetics of Aβ 1-42 aggregation by DDX3X seeds. B . Boltzmann fitting of the cross-seeding experiment depicting the co-aggregation kinetics of Aβ 1-42 when cross-seeded with sonicated seeds of DDX3X variants. C . Representative confocal images of N2a cells expressing WT or mutant DDX3X-mCherry, subjected to SA stress (100µM), wash-off, and recovery, followed by ThT staining and immunostained with DAPI (blue). ThT fluorescence is represented in green. Scale bar= 10µm. D-F . Quantification of numbers of DDX3X (red), ThT stained (green) and ThT-DDX3X (yellow) granules per cell from ≥ 20 cells across three independent experiments exhibiting DDX3X colocalization with ThT granules in N2a cells. ****p < 0.0001 (DDX3X granules/cell), ****p < 0.0001, ***p = 0.0005, **p = 0.0022 (WT vs I190S), **p = 0.0080 (WT vs T198P) (ThT granules/cells) ****p < 0.0001, ***p = 0.0003, **p = 0.0021 (WT vs I190S), **p = 0.0057 (WT vs T198P) (ThT-DDX3X colocalised granules/cells) (One-way ANOVA test). Data shown are mean ± SEM.

Article Snippet: Full length DDX3X-mCherry construct (pReceiver-M56-DDX3X-mCherry ) was procured from GeneCopoeia.

Techniques: Sonication, Expressing, Mutagenesis, Staining, Fluorescence

Journal: bioRxiv

Article Title: Keratin 5 marks cancer-propagating cells sustained by an osteopontin-producing niche in high-grade serous ovarian carcinoma

doi: 10.64898/2026.01.28.702332

Figure Lengend Snippet: A , Structure of Lenti-KRT5mCherry (L-KRT5mCherry). B , Co-localization of KRT5 immunostaining (KRT5, green) and mCherry (L-KRT5) expression (magenta). Orange, overlay. Counterstaining with DAPI (blue). Scale bar, 60 µm. C , Quantification of cells expressing both KRT5 and L-KRT5 (KRT5+ L-KRT5+), or only KRT5 (KRT5+) or L-KRT5+. D , Experimental design of isolation of cells expressing both L-KRT5mCherry and L-hUbC-GFP or L-hUbC-GFP alone. E, PCR detection of L-hUbC-GFP (GFP) and L-KRT5mCherry (mCherry) DNA in both KRT5+ (K5+) and KRT5- (K5-) cells. All error bars denote s.d.

Article Snippet: Briefly, for lentivirus packaging psPAX2 (Addgene, 12260), pMD2.G envelope plasmid (Addgene, 12259), and KRT5 promoter clone (GeneCopoeia, HRPM15909-LvPM02, mCherry) were employed.

Techniques: Immunostaining, Expressing, Isolation

Journal: bioRxiv

Article Title: Keratin 5 marks cancer-propagating cells sustained by an osteopontin-producing niche in high-grade serous ovarian carcinoma

doi: 10.64898/2026.01.28.702332

Figure Lengend Snippet: A and B , Organoids derived from SKOV3 cells expressing both lentiviruses (A, GFP and mCherry, orange) or GFP alone (B, green). Scale bar, 100 µm. C , Quantification of KRT5+ (blue symbols, pink bars) and KRT5- (pink symbols, yellow bars) cancer organoids in 6 consecutive passages. All error bars denote s.d. D, Volume of tumors formed by serially diluted (1 x 10 5 , 1 x 10 4 , 1 x 10 3 ) of KRT5+ and KRT5- cells after their s.c. transplantation into different flanks of NSG mice. KRT5-group did not form tumors. E and F , mCherry (E) and KRT5 (F) expression in KRT5+ cell derived xenografts. Elite ABC method. Hematoxylin counterstaining. Scale bar, 60 µm. All error bars denote s.d. G. Live microscopy of cells were isolated by FACS based on their expression of GFP (green) and mCherry (magenta) after coinfection with Lenti-UbC-GFP and Lenti-KRT5mCherry. Orange, Overlay. Individual frames of live microscopy. Scale bar, 60 µm.

Article Snippet: Briefly, for lentivirus packaging psPAX2 (Addgene, 12260), pMD2.G envelope plasmid (Addgene, 12259), and KRT5 promoter clone (GeneCopoeia, HRPM15909-LvPM02, mCherry) were employed.

Techniques: Derivative Assay, Expressing, Transplantation Assay, Microscopy, Isolation

Journal: bioRxiv

Article Title: Disruption of ADAMTSL4 Causes Ectopia Pupillae in Zebrafish via COL8A1-Driven Cell Migration

doi: 10.1101/2025.11.03.686242

Figure Lengend Snippet: (A). qPCR analysis showing fold changes in mRNA expression of ECM-associated genes in EP eyes of adamtsl4 ⁻/⁻ zebrafish mutants compared with normal eyes of sibling controls. Each group included three biological replicates. (B). qPCR validation of ECM-associated gene expression in human RPEs following siRNA-mediated knockdown of ADAMTSL4 , compared to negative siRNA controls. Each group has three biological replications. (C). Western blot analysis of ADAMTSL4, TGFB2, and COL8A1 protein levels in ADAMTSL4 knockdown RPEs versus controls. GAPDH was used as a loading control. Each group contained three biological replicates. (D). Co-IP between ADAMTSL4 and COL8A1 in HEK293T cells. Exogenous ADAMTSL4 fused with EGFP and COL8A1 fused with mCherry were expressed and precipitated using EGFP antibody-coated beads. Interacting proteins were detected by Western blot. (E). Knockdown of COL8A1 in ADAMTSL4- deficient RPE cells using siRNA. Knockdown efficiency was confirmed by qPCR. (F). Wound healing assay showing significantly reduced cell migration in RPEs following ADAMTSL4 knockdown, COL8A1 knockdown, combined knockdown. Negative siRNA control was transfected toaccount for potential cytotoxic effects of transfection, and total siRNA concentrations were maintained consistently across all groups. Quantification was based on the percentage change in wound area across at least three independent experiments. Statistical significance is indicated by asterisks: ns P ≥ 0.05, * P < 0.05, ** P < 0.01, *** P < 0.001, **** P < 0.0001; error bars indicate the standard error of the mean. Co-IP, Co-Immunoprecipitation; qPCR, quantitative polymerase chain reaction; siRNA, small interfering RNA; RPE, retinal pigmented epithelium; HEK293T, Human Embryonic Kidney 293T cell; IP, immunoprecipitation; IB, immunoblotting.

Article Snippet: The primary antibodies used for immunoblotting were as follows: ADAMTSL4 Rabbit pAb (Abclonal, #A4785, 1:1000), TGFB2 Rabbit pAb (Abclonal, #A3640, 1:1000 dilution), Collagen VIII alpha 1 Rabbit pAb (Affinity, #DF8902,1:1000 dilution), GFP tag Mouse mAb (Proteintech, #66002-1-lg, 1:20000 dilution), mCherry Rabbit pAb (Proteintech, #26765-1-AP, 1:2000), and GAPDH Rabbit pAb (Proteintech, #10494-1-AP, 1:5000 dilution).

Techniques: Expressing, Biomarker Discovery, Gene Expression, Knockdown, Western Blot, Control, Co-Immunoprecipitation Assay, Wound Healing Assay, Migration, Transfection, Immunoprecipitation, Real-time Polymerase Chain Reaction, Small Interfering RNA