Journal: Experimental Animals

Article Title: Ubiquitin-like 4A alleviates the progression of intracerebral hemorrhage by regulating oxidative stress and mitochondrial damage

doi: 10.1538/expanim.24-0035

Figure Lengend Snippet: (a) Schematic diagram showing the experimental procedure of the ICH rat. (b) Forelimb placing test and corner turn test of rats. (c) Brain water content of rats. (d) Real-time PCR and western blot were used to verify the expression of UBL4A in brain tissue. (e) Typical hematoxylin-eosin (HE) staining images presenting the striatal hematomas in rats. Scale bar: 500 µ m (left) and 100 µ m (right). (f) Fluoro-Jade B (FJB) staining was performed to detect neuronal cell death in rats. Scale bar: 50 µ m. (g) Co-localization of ubiquitin-like protein 4A (UBL4A) and NeuN was detected by immunofluorescence staining. Scale bar: 50 µ m. ICH, intracerebral hemorrhage. * P <0.05, ** P <0.01, ns: no significance.

Article Snippet: Tissue sections were blocked with UBL4A (1:100; 14253-1-AP; Proteintech, Wuhan, China) and Neuronal Nucleus antibody (NeuN; 1:300; ab104224; Abcam, Shanghai, China) overnight at 4°C, followed by FITC-labeled goat anti-rabbit IgG (1:200; ab6717; Abcam) and Cy3-labeled goat anti-mouse IgG (1:200; A-21424; Invitrogen, Carlsbad, CA, USA) were applied in a dark box for 90 min. After rinsing with PBS, 4’,6-diamidino-2-phenylindole (DAPI; Aladdin, Shanghai, China) was used as a counterstain.

Techniques: Real-time Polymerase Chain Reaction, Western Blot, Expressing, Staining, Ubiquitin Proteomics, Immunofluorescence

Journal: Experimental Animals

Article Title: Ubiquitin-like 4A alleviates the progression of intracerebral hemorrhage by regulating oxidative stress and mitochondrial damage

doi: 10.1538/expanim.24-0035

Figure Lengend Snippet: (a) The diagram depicts the flow chart of the cell experiment. (b) Real-time PCR and western blot were used to verify the expression level of ubiquitin-like protein 4A (UBL4A) in neurons. (c) CCK-8 assay was performed to determine cell viability. (d) TUNEL staining was used to detect cell apoptosis. Scale bar: 50 µ m. LV-shNC, negative control shRNA; LV-shUBL4A, shUBL4A knockdown; LV-vector, empty vector; LV-UBL4A, UBL4A overexpression. * P <0.05, ** P <0.01.

Article Snippet: Tissue sections were blocked with UBL4A (1:100; 14253-1-AP; Proteintech, Wuhan, China) and Neuronal Nucleus antibody (NeuN; 1:300; ab104224; Abcam, Shanghai, China) overnight at 4°C, followed by FITC-labeled goat anti-rabbit IgG (1:200; ab6717; Abcam) and Cy3-labeled goat anti-mouse IgG (1:200; A-21424; Invitrogen, Carlsbad, CA, USA) were applied in a dark box for 90 min. After rinsing with PBS, 4’,6-diamidino-2-phenylindole (DAPI; Aladdin, Shanghai, China) was used as a counterstain.

Techniques: Real-time Polymerase Chain Reaction, Western Blot, Expressing, Ubiquitin Proteomics, CCK-8 Assay, TUNEL Assay, Staining, Negative Control, shRNA, Knockdown, Plasmid Preparation, Over Expression

Journal: Experimental Animals

Article Title: Ubiquitin-like 4A alleviates the progression of intracerebral hemorrhage by regulating oxidative stress and mitochondrial damage

doi: 10.1538/expanim.24-0035

Figure Lengend Snippet: (a) The level of reactive oxygen species (ROS) in cells was determined by flow cytometry using a dichlorodihydrofluorescein diacetate (DCFH-DA) based kit (cell number: ×10 3 ). (b) Manganese superoxide dismutase (MnSOD) activity was measured with MnSOD assay kit. (c) Detection of mitochondrial membrane potential by JC-1 staining. (d) The relative mitochondrial DNA (mtDNA) level was detected by real-time PCR. LV-shNC, negative control shRNA; LV-shUBL4A, shUBL4A knockdown; LV-vector, empty vector; LV-UBL4A, UBL4A overexpression. * P <0.05, ** P <0.01.

Article Snippet: Tissue sections were blocked with UBL4A (1:100; 14253-1-AP; Proteintech, Wuhan, China) and Neuronal Nucleus antibody (NeuN; 1:300; ab104224; Abcam, Shanghai, China) overnight at 4°C, followed by FITC-labeled goat anti-rabbit IgG (1:200; ab6717; Abcam) and Cy3-labeled goat anti-mouse IgG (1:200; A-21424; Invitrogen, Carlsbad, CA, USA) were applied in a dark box for 90 min. After rinsing with PBS, 4’,6-diamidino-2-phenylindole (DAPI; Aladdin, Shanghai, China) was used as a counterstain.

Techniques: Flow Cytometry, Activity Assay, Membrane, Staining, Real-time Polymerase Chain Reaction, Negative Control, shRNA, Knockdown, Plasmid Preparation, Over Expression

Journal: Experimental Animals

Article Title: Ubiquitin-like 4A alleviates the progression of intracerebral hemorrhage by regulating oxidative stress and mitochondrial damage

doi: 10.1538/expanim.24-0035

Figure Lengend Snippet: (a) Real-time PCR to verify the expression of miR-34a-5p in neurons. (b) Real-time PCR and western blot were used to verify the expression level of ubiquitin-like protein 4A (UBL4A) in neurons with miR-34a-5p knockdown. (c) TUNEL staining was used to detect cell apoptosis. Scale bar: 50 µ m. (d) The level of reactive oxygen species (ROS) in cells was determined by flow cytometry using a dichlorodihydrofluorescein diacetate (DCFH-DA) based kit (cell number: ×10 3 ). LV-shNC, negative control shRNA; LV-miR-34a-sponge, miR-34a-5p knockdown. ** P <0.01.

Article Snippet: Tissue sections were blocked with UBL4A (1:100; 14253-1-AP; Proteintech, Wuhan, China) and Neuronal Nucleus antibody (NeuN; 1:300; ab104224; Abcam, Shanghai, China) overnight at 4°C, followed by FITC-labeled goat anti-rabbit IgG (1:200; ab6717; Abcam) and Cy3-labeled goat anti-mouse IgG (1:200; A-21424; Invitrogen, Carlsbad, CA, USA) were applied in a dark box for 90 min. After rinsing with PBS, 4’,6-diamidino-2-phenylindole (DAPI; Aladdin, Shanghai, China) was used as a counterstain.

Techniques: Real-time Polymerase Chain Reaction, Expressing, Western Blot, Ubiquitin Proteomics, Knockdown, TUNEL Assay, Staining, Flow Cytometry, Negative Control, shRNA

Journal: Experimental Animals

Article Title: Ubiquitin-like 4A alleviates the progression of intracerebral hemorrhage by regulating oxidative stress and mitochondrial damage

doi: 10.1538/expanim.24-0035

Figure Lengend Snippet: (a) Predicted miR-34a-5p target sequences in 3′-UTR of Ubl4a (Left). Dual-luciferase reporter assay was conducted to verify whether ubiquitin-like protein 4A (UBL4A) was the target of miR-34a-5p (Right). NC mimics, negative control mimics; WT, wild type; MUT, mutant type. (b) Western blot was used to verify the expression level of UBL4A in neurons. (c) TUNEL staining was used to detect cell apoptosis. Scale bar: 50 µ m. (d) The level of reactive oxygen species (ROS) in cells was determined by flow cytometry using a dichlorodihydrofluorescein diacetate (DCFH-DA) based kit (cell number: ×10 3 ). LV-shNC, negative control shRNA; LV-miR-34a-sponge, miR-34a-5p knockdown. LV-shUBL4A, shUBL4A knockdown. * P <0.05, ** P <0.01.

Article Snippet: Tissue sections were blocked with UBL4A (1:100; 14253-1-AP; Proteintech, Wuhan, China) and Neuronal Nucleus antibody (NeuN; 1:300; ab104224; Abcam, Shanghai, China) overnight at 4°C, followed by FITC-labeled goat anti-rabbit IgG (1:200; ab6717; Abcam) and Cy3-labeled goat anti-mouse IgG (1:200; A-21424; Invitrogen, Carlsbad, CA, USA) were applied in a dark box for 90 min. After rinsing with PBS, 4’,6-diamidino-2-phenylindole (DAPI; Aladdin, Shanghai, China) was used as a counterstain.

Techniques: Luciferase, Reporter Assay, Ubiquitin Proteomics, Negative Control, Mutagenesis, Western Blot, Expressing, TUNEL Assay, Staining, Flow Cytometry, shRNA, Knockdown

Journal: Computational and Structural Biotechnology Journal

Article Title: Identification and validation of protein biomarkers for predicting gastrointestinal stromal tumor recurrence

doi: 10.1016/j.csbj.2024.02.017

Figure Lengend Snippet: The clinical and pathological characteristics of patients with and without recurrence.

Article Snippet: The information of primary antibodies is listed as follows: GPX4 (Proteintech 67763–1-lg), RBM4 (Affinity DF12225), TPM3 (Affinity DF6338), PFKFB2 (Immunoway YP1872), PGAM5 (Proteintech 28445–1-AP).

Techniques:

Journal: Computational and Structural Biotechnology Journal

Article Title: Identification and validation of protein biomarkers for predicting gastrointestinal stromal tumor recurrence

doi: 10.1016/j.csbj.2024.02.017

Figure Lengend Snippet: Expressions of GPX4, RBM4, TPM3, PFKFB2, PGAM5 in primary GIST tissues from 23 recurrent and 25 non-recurrent patients. Representative IHC staining (100X and 400X) and quantification of five random fields by mean of integrated optical density (IOD) using Image-Pro Plus in different markers and groups. (* P < 0.05, ** P < 0.01, *** P < 0.001).

Article Snippet: The information of primary antibodies is listed as follows: GPX4 (Proteintech 67763–1-lg), RBM4 (Affinity DF12225), TPM3 (Affinity DF6338), PFKFB2 (Immunoway YP1872), PGAM5 (Proteintech 28445–1-AP).

Techniques: Immunohistochemistry

Journal: Computational and Structural Biotechnology Journal

Article Title: Identification and validation of protein biomarkers for predicting gastrointestinal stromal tumor recurrence

doi: 10.1016/j.csbj.2024.02.017

Figure Lengend Snippet: The prognostic value of GPX4, RBM4, TPM3, PFKFB2, PGAM5 in 48 patients with GIST. Comparison of Recurrence-free survival (RFS) and overall survival (OS) between patients with high expressions of these five proteins and low expressing cases in GIST via Kaplan–Meier analysis. (* P < 0.05, ** P < 0.01, *** P < 0.001).

Article Snippet: The information of primary antibodies is listed as follows: GPX4 (Proteintech 67763–1-lg), RBM4 (Affinity DF12225), TPM3 (Affinity DF6338), PFKFB2 (Immunoway YP1872), PGAM5 (Proteintech 28445–1-AP).

Techniques: Comparison, Expressing

Journal: Computational and Structural Biotechnology Journal

Article Title: Identification and validation of protein biomarkers for predicting gastrointestinal stromal tumor recurrence

doi: 10.1016/j.csbj.2024.02.017

Figure Lengend Snippet: Correlations between GPX4/RBM4/TPM3/PFKFB2/PGAM5 expressions and clinical features.

Article Snippet: The information of primary antibodies is listed as follows: GPX4 (Proteintech 67763–1-lg), RBM4 (Affinity DF12225), TPM3 (Affinity DF6338), PFKFB2 (Immunoway YP1872), PGAM5 (Proteintech 28445–1-AP).

Techniques:

Journal: Research in Veterinary Science

Article Title: EIF4A2 interacts with the membrane protein of transmissible gastroenteritis coronavirus and plays a role in virus replication

doi: 10.1016/j.rvsc.2018.12.005

Figure Lengend Snippet: Primers for QRT-PCR.

Article Snippet: The beta tubulin rabbit polyclonal antibody (β-tubulin) and EIF4A2 rabbit polyclonal antibody were purchased from Proteintech (USA).

Techniques: Sequencing

Journal: Research in Veterinary Science

Article Title: EIF4A2 interacts with the membrane protein of transmissible gastroenteritis coronavirus and plays a role in virus replication

doi: 10.1016/j.rvsc.2018.12.005

Figure Lengend Snippet: GST-pull down and CO-IP were used to analyze the interaction between the M protein and candidate protein EIF4A2 by SDS-PAGE and Western blot. (A) The results of the GST pull-down by SDS-PAGE M: Prestained protein marker; 1: purified GST-M fusion protein + purified His-EIF4A2 fusion protein; 2: purified GST-tag protein + purified His-EIF4A2 fusion protein; 3: purified His-EIF4A2 fusion protein; 4: purified GST-M fusion protein; 5: purified GST-tag protein; (B) The results of the GST pull-down by Western blot M: Prestained protein marker; 1–2: His-EIF4A2 fusion protein. (C) pIECs infected with TGEV and control cells were harvested at 24 h, and the M protein was precipitated by an antibody in TGEV-infected group but not the mock group. TGEV+ represent the pIECs infected with TGEV and TGEV- represent uninfected TGEV in pIECs.

Article Snippet: The beta tubulin rabbit polyclonal antibody (β-tubulin) and EIF4A2 rabbit polyclonal antibody were purchased from Proteintech (USA).

Techniques: Co-Immunoprecipitation Assay, SDS Page, Western Blot, Marker, Purification, Infection, Control

Journal: Research in Veterinary Science

Article Title: EIF4A2 interacts with the membrane protein of transmissible gastroenteritis coronavirus and plays a role in virus replication

doi: 10.1016/j.rvsc.2018.12.005

Figure Lengend Snippet: Co-localization of EIF4A2 with the M protein. Determining the location of the M protein using FITC-conjugated goat anti-rat IgG (green) as the secondary antibody after pIECs were infected with TGEV. The red colour indicates the distribution domain of EIF4A2. The nucleus was stained with DAPI (blue). A ZEISS LSM 800 with Airyscan was used to examine the triple-stained location. (For interpretation of the references to colour in this figure legend, the reader is referred to the web version of this article.)

Article Snippet: The beta tubulin rabbit polyclonal antibody (β-tubulin) and EIF4A2 rabbit polyclonal antibody were purchased from Proteintech (USA).

Techniques: Infection, Staining

Journal: Research in Veterinary Science

Article Title: EIF4A2 interacts with the membrane protein of transmissible gastroenteritis coronavirus and plays a role in virus replication

doi: 10.1016/j.rvsc.2018.12.005

Figure Lengend Snippet: EIF4A2 gene silencing. pIECs were cultured in six-well culture plates and transfected with siRNAs (20 nM). The cell lysates were harvested for Western blotting at 48 h. The average densitometric intensity of EIF4A2 in the immunoblot analysis, with β-tubulin used as a control (A and B). The relativelevel of EIF4A2 mRNA expression was used to further evaluate the interference effect of siRNA2 by RT-qPCR at 36 h (C). *P < 0.05.

Article Snippet: The beta tubulin rabbit polyclonal antibody (β-tubulin) and EIF4A2 rabbit polyclonal antibody were purchased from Proteintech (USA).

Techniques: Cell Culture, Transfection, Western Blot, Control, Expressing, Quantitative RT-PCR

Journal: Research in Veterinary Science

Article Title: EIF4A2 interacts with the membrane protein of transmissible gastroenteritis coronavirus and plays a role in virus replication

doi: 10.1016/j.rvsc.2018.12.005

Figure Lengend Snippet: Virus titration. EIF4A2-knockdown with siRNA2 cells and negative control knockdown cells were adsorbed with TGEV (MOI = 0.1) at 37 °C for 1 h. The culture supernatants of the cells and the virus-associated cells infected with TGEV were collected at different points. The viral titers are presented as the averages from three independent samples. **P < 0.01 and ***P < 0.001.

Article Snippet: The beta tubulin rabbit polyclonal antibody (β-tubulin) and EIF4A2 rabbit polyclonal antibody were purchased from Proteintech (USA).

Techniques: Virus, Titration, Knockdown, Negative Control, Infection

Journal: bioRxiv

Article Title: Intraflagellar transport protein 74 is essential for mouse spermatogenesis and male fertility by regulating axonemal microtubule assembly in mice

doi: 10.1101/457804

Figure Lengend Snippet: A.Examination of selective IFT protein expression levels in the control and conditional Ift74 knockout mice by Western blot. Compared to the controls, the expression levels of IFT27, IFT57, IFT81, IFT88 and IFT140, but not IFT20 and IFT25 are reduced in the conditional Ift74 knockout mice. a: representative Western blot result; b: quantitative analysis of selective IFT protein expression. B. Examination of testicular expression levels of ODF2, a component of sperm tail outer dense fibers, AKAP4, a major component of fibrous sheath, and SPAG16L, a component of axonemal central apparatus protein in the control and conditional Ift74 knockout mice. There was no difference in ODF2 and SPAG16L expression. However, the processed AKAP4 was significantly reduced in the conditional Ift74 knockout mice. β-ACTIN was used as controls. a: representative Western blot result; b: quantitative analysis of ODF2 and SPAG16L; c: quantitative analysis of AKAP4.

Article Snippet: Then the Nonspecific sites were blocked in a Tris-buffered saline solution containing 5% nonfat dry milk powder and 0.05% Tween 20 (TBST) for 1 hour at room temperature, and the membranes were incubated with indicated primary antibodies (IFT25: 1:2000, Cat No: 15732-1-AP, ProteinTech; IFT74: 1:2000, Cat No: AAS27620e from ANTIBODY VERIFY; IFT81: 1:1000, Cat No: 11744-1-AP, ProteinTech; β-actin: 1:2000, Cat No: 4967 S, Cell Signaling Antibodies against IFT20, IFT27, IFT57, IFT88 and IFT140 (1:2000) were from Dr. Pazour’s laboratory ( , , ); AKAP4: 1:4000, from Dr. George Gerton at University of Pennsylvania; ODF2: 1:800, Cat No: 12058-1-AP, ProteinTech; SPAG16L: 1:1000, generated by Z.Z.’s laboratory) at 4°C overnight.

Techniques: Expressing, Knock-Out, Western Blot