Journal: Animals : an Open Access Journal from MDPI

Article Title: SPARCL1 Influences Bovine Skeletal Muscle-Derived Satellite Cell Migration and Differentiation through an ITGB1-Mediated Signaling Pathway

doi: 10.3390/ani10081361

Figure Lengend Snippet: Secreted protein acidic and rich in cysteine like 1 (SPARCL1) interacts with integrin β1 (ITGB1). ( A ) Co-immunoprecipitation with an anti-SPARCL1 antibody followed by Western blotting with an anti-ITGB1 antibody. ( B ) Co-immunoprecipitation with an anti-ITGB1 antibody followed by Western blotting with an anti-SPARCL1 antibody. Input, IgG, and IP represent the positive control, negative control, and target experimental group, respectively.

Article Snippet: The following primary antibodies were used: anti-SPARCL1, anti-ITGB1, anti-p-FAK, anti-FAK, anti-p-paxillin, anti-paxillin, anti-vinculin, anti-Arp2/3, anti-Cdc42, anti-MyoD, and anti-GAPDH (all at dilution 1:500; Bioss Antibodies, China).

Techniques: Immunoprecipitation, Western Blot, Positive Control, Negative Control

Journal: Animals : an Open Access Journal from MDPI

Article Title: SPARCL1 Influences Bovine Skeletal Muscle-Derived Satellite Cell Migration and Differentiation through an ITGB1-Mediated Signaling Pathway

doi: 10.3390/ani10081361

Figure Lengend Snippet: SPARCL1 influences MDSC migration and differentiation. ( A , C ) Cell scratch assay images at 20 h after SPARCL1 activation or inhibition. ( B , D ) Quantification of the cell migration rate at 20 h after SPARCL1 activation or inhibition according to ( A , C ). ( E , G ) Desmin immunofluorescence staining in MDSCs after SPARCL1 activation or inhibition. ( F , H ) Quantification of the myotube rate based on ( E , G ). ( I , K ) Western blotting images of MyoD expression after SPARCL1 activation or inhibition. ( J , L ) Quantification of MyoD from Western blotting results presented in ( I , K ). Scale bar = 100 μm. * p < 0.05, ** p < 0.01, *** p < 0.001.

Article Snippet: The following primary antibodies were used: anti-SPARCL1, anti-ITGB1, anti-p-FAK, anti-FAK, anti-p-paxillin, anti-paxillin, anti-vinculin, anti-Arp2/3, anti-Cdc42, anti-MyoD, and anti-GAPDH (all at dilution 1:500; Bioss Antibodies, China).

Techniques: Migration, Wound Healing Assay, Activation Assay, Inhibition, Immunofluorescence, Staining, Western Blot, Expressing

Journal: Animals : an Open Access Journal from MDPI

Article Title: SPARCL1 Influences Bovine Skeletal Muscle-Derived Satellite Cell Migration and Differentiation through an ITGB1-Mediated Signaling Pathway

doi: 10.3390/ani10081361

Figure Lengend Snippet: SPARCL1 affects an ITGB1-mediated signaling pathway. ( A , I ) Western blotting results for ITGB1, p-FAK, FAK, p-paxillin, paxillin, vinculin, Cdc42, and Arp2/3 after SPARCL1 activation or inhibition, respectively. ( B – H ) Quantification of the Western blotting results presented in ( A ). ( J – P ) Quantification of the Western blotting results presented in ( I ). ** p < 0.01.

Article Snippet: The following primary antibodies were used: anti-SPARCL1, anti-ITGB1, anti-p-FAK, anti-FAK, anti-p-paxillin, anti-paxillin, anti-vinculin, anti-Arp2/3, anti-Cdc42, anti-MyoD, and anti-GAPDH (all at dilution 1:500; Bioss Antibodies, China).

Techniques: Western Blot, Activation Assay, Inhibition

Journal: Animals : an Open Access Journal from MDPI

Article Title: SPARCL1 Influences Bovine Skeletal Muscle-Derived Satellite Cell Migration and Differentiation through an ITGB1-Mediated Signaling Pathway

doi: 10.3390/ani10081361

Figure Lengend Snippet: SPARCL1 influences cell migration and differentiation through ITGB1. ( A ) Cell scratch assay images after simultaneous SPARCL1 activation and ITGB1 inhibition. ( B ) Quantification of the cell migration rate based on ( A ). ( C ) Desmin immunofluorescence staining of MDSCs after SPARCL1 activation and ITGB1 inhibition. ( D ) Quantification of the myotube rate represented in ( C ). ( E ) Western blotting results of MyoD expression after SPARCL1 activation and ITGB1 inhibition. ( F ) Quantification of the MyoD Western blotting results presented in ( E ). pSPgRNA is the blank control for the SPARCL1 activation group, NC is the negative control for the ITGB1 siRNA interference group, VPR-S represents the SPARCL1 activation group, and siRNA-I represents the ITGB1 siRNA interference group. * p < 0.05, ** p < 0.01, *** p < 0.001; scale bar = 100 μm.

Article Snippet: The following primary antibodies were used: anti-SPARCL1, anti-ITGB1, anti-p-FAK, anti-FAK, anti-p-paxillin, anti-paxillin, anti-vinculin, anti-Arp2/3, anti-Cdc42, anti-MyoD, and anti-GAPDH (all at dilution 1:500; Bioss Antibodies, China).

Techniques: Migration, Wound Healing Assay, Activation Assay, Inhibition, Immunofluorescence, Staining, Western Blot, Expressing, Negative Control

Journal: Animals : an Open Access Journal from MDPI

Article Title: SPARCL1 Influences Bovine Skeletal Muscle-Derived Satellite Cell Migration and Differentiation through an ITGB1-Mediated Signaling Pathway

doi: 10.3390/ani10081361

Figure Lengend Snippet: SPARCL1 regulates an ITGB1-mediated signaling pathway through ITGB1. ( A ) Western blotting results for ITGB1, p-FAK, FAK, p-paxillin, paxillin, vinculin, Cdc42, and Arp2/3 expression after SPARCL1 activation and ITGB1 inhibition. ( B – H ) Quantification of the Western blotting results presented in ( A ). pSPgRNA is the blank control for the SPARCL1 activation group, NC is the negative control for the ITGB1 siRNA interference group, VPR-S represents the SPARCL1 activation group, and siRNA-I represents the ITGB1 siRNA interference group. * p < 0.05, ** p < 0.01, *** p < 0.001.

Article Snippet: The following primary antibodies were used: anti-SPARCL1, anti-ITGB1, anti-p-FAK, anti-FAK, anti-p-paxillin, anti-paxillin, anti-vinculin, anti-Arp2/3, anti-Cdc42, anti-MyoD, and anti-GAPDH (all at dilution 1:500; Bioss Antibodies, China).

Techniques: Western Blot, Expressing, Activation Assay, Inhibition, Negative Control

Journal: Animals : an Open Access Journal from MDPI

Article Title: SPARCL1 Influences Bovine Skeletal Muscle-Derived Satellite Cell Migration and Differentiation through an ITGB1-Mediated Signaling Pathway

doi: 10.3390/ani10081361

Figure Lengend Snippet: Proposed model for SPARCL1 regulation of bovine MDSC migration and differentiation through ITGB1. As an extracellular matrix protein, SPARCL1 interacts with the transmembrane receptor ITGB1 and regulates changes in the expression of adhesion-plaque-associated proteins, such as ITGB1, p-FAK, FAK, p-paxillin, paxillin, and vinculin to affect the formation of focal adhesions. The Cdc42 and Arp2/3 complex was also affected by SPARCL1 through ITGB1. Further, bovine MDSC migration was stimulated by SPARCL1, and cells gathered and apparently expressed MyoD to begin an early-stage differentiation.

Article Snippet: The following primary antibodies were used: anti-SPARCL1, anti-ITGB1, anti-p-FAK, anti-FAK, anti-p-paxillin, anti-paxillin, anti-vinculin, anti-Arp2/3, anti-Cdc42, anti-MyoD, and anti-GAPDH (all at dilution 1:500; Bioss Antibodies, China).

Techniques: Migration, Expressing

Journal: Journal of Orthopaedic Translation

Article Title: SPARCL1 promotes chondrocytes extracellular matrix degradation and inflammation in osteoarthritis via TNF/NF-κB pathway

doi: 10.1016/j.jot.2024.02.009

Figure Lengend Snippet: The expression level of SPARCL1 in human OA cartilage. A Microarray profiling analysis data of SPARCL1 expression level in OA cartilage and normal cartilage. Date was originated from GSE113825. Unpaired T-test were used. B Immunohistochemistry assay with anti-SPARCL1 in undamaged and OA cartilage tissues. Scale bar, Left, 100 μm; Right, 50 μm. C Relative SPARCL1 expression level in OA (n = 55) and corresponding undamaged (n = 55) cartilage tissues based on an immunohistochemistry assay and significance was evaluated by paired Student's t test. Data are expressed as mean ± SD, *P < 0.05; **P < 0.01; ***P < 0.001.

Article Snippet: Proteins were analyzed with antibodies against matrix metalloproteinases 3 (MMP3) (1:1000, Santa Cruz Cat# sc-21732), ADAMTS5 (1:1 000, Affinity Cat# DF13268), FAK (1:1000, Cell Signaling Technology Cat# 3285), Phospho-FAK (1:1000, Cell Signaling Technology Cat# 3285), NFκB (1:1000, Cell Signaling Technology Cat# 6956), IKKα (1:1000, Cell Signaling Technology Cat# 2682), IKKβ (1:1000, Cell Signaling Technology Cat# 2370), SPARCL1 (1:1000, Proteintech Cat#13517-1-AP), β-ACTIN (1:1000, Servicebio Cat# GB11001), Goat Anti-Rabbit IgG (1:5000, Jackson Cat#111–005) and Goat Anti-mouse IgG (1:5000, Jackson Cat#115–005).

Techniques: Expressing, Microarray, Immunohistochemistry

Journal: Journal of Orthopaedic Translation

Article Title: SPARCL1 promotes chondrocytes extracellular matrix degradation and inflammation in osteoarthritis via TNF/NF-κB pathway

doi: 10.1016/j.jot.2024.02.009

Figure Lengend Snippet: Sparcl1 regulated ECM degradation in chondrocytes. A Alcian blue staining of ATDC5 with micromass culture in chondrogenesis media with or without recombinant Sparcl1 protein. Representative image of staining at day 8 is shown. B CCK-8 assay test of the cell viability of MCC treated with different concentration of recombinant Sparcl1 protein (10, 25, 50, 100 ng/ml). C Quantification of mRNA levels for Col2a1, Acan and Sox9 in MCC treated with recombinant Sparcl1 protein (10 and 50 ng/ml) for 48 h. D Quantification of mRNA levels for Mmp3, Mmp13, Adamts1, Adamts5 in MCC treated with recombinant Sparcl1 protein (10 and 50 ng/ml) for 48 h. E Quantification of mRNA levels for Col2a1, Acan, Mmp3, Mmp13, Ccl2 and IL-6 in MCC treated with IL-1β (10 ng/ml) and recombinant Sparcl1 protein (10, 25 and 50 ng/ml) for 48 h. Data are expressed as mean ± SD, *P < 0.05; **P < 0.01; ***P < 0.001. One-way ANOVA with post-hoc Bonferroni correction was used for comparisons.

Article Snippet: Proteins were analyzed with antibodies against matrix metalloproteinases 3 (MMP3) (1:1000, Santa Cruz Cat# sc-21732), ADAMTS5 (1:1 000, Affinity Cat# DF13268), FAK (1:1000, Cell Signaling Technology Cat# 3285), Phospho-FAK (1:1000, Cell Signaling Technology Cat# 3285), NFκB (1:1000, Cell Signaling Technology Cat# 6956), IKKα (1:1000, Cell Signaling Technology Cat# 2682), IKKβ (1:1000, Cell Signaling Technology Cat# 2370), SPARCL1 (1:1000, Proteintech Cat#13517-1-AP), β-ACTIN (1:1000, Servicebio Cat# GB11001), Goat Anti-Rabbit IgG (1:5000, Jackson Cat#111–005) and Goat Anti-mouse IgG (1:5000, Jackson Cat#115–005).

Techniques: Staining, Recombinant, CCK-8 Assay, Concentration Assay

Journal: Journal of Orthopaedic Translation

Article Title: SPARCL1 promotes chondrocytes extracellular matrix degradation and inflammation in osteoarthritis via TNF/NF-κB pathway

doi: 10.1016/j.jot.2024.02.009

Figure Lengend Snippet: Sparcl1 accelerated OA process in ACLT mice model. A Representative safranin O-fast green images of osteoarthritic knee joints, which were collected 6 weeks after ACLT surgery. Yellow arrowheads indicated articular cartilage degradation. n = 8; Scale bar, left, 200 μm; right, 100 μm. B The severity of OA-like phenotype was analyzed using the Osteoarthritis Research Society International (OARSI) score system (n = 8). C The degree of synovitis was semi-quantified by the enlargement of synovial lining layers and density of cells (n = 8). D Three-dimensional models of mice knee joints. Red arrow showed osteophyte formation. Scale bar, 250 μm. E, F Osteophytes were semi-quantified by evaluating the osteophyte formation score consisting of two domains, size (E) and maturity (F) (n = 5) Data are expressed as mean ± SD, *P < 0.05; **P < 0.01; ***P < 0.001. One-way ANOVA with post-hoc Bonferroni correction was used for comparisons.

Article Snippet: Proteins were analyzed with antibodies against matrix metalloproteinases 3 (MMP3) (1:1000, Santa Cruz Cat# sc-21732), ADAMTS5 (1:1 000, Affinity Cat# DF13268), FAK (1:1000, Cell Signaling Technology Cat# 3285), Phospho-FAK (1:1000, Cell Signaling Technology Cat# 3285), NFκB (1:1000, Cell Signaling Technology Cat# 6956), IKKα (1:1000, Cell Signaling Technology Cat# 2682), IKKβ (1:1000, Cell Signaling Technology Cat# 2370), SPARCL1 (1:1000, Proteintech Cat#13517-1-AP), β-ACTIN (1:1000, Servicebio Cat# GB11001), Goat Anti-Rabbit IgG (1:5000, Jackson Cat#111–005) and Goat Anti-mouse IgG (1:5000, Jackson Cat#115–005).

Techniques:

Journal: Journal of Orthopaedic Translation

Article Title: SPARCL1 promotes chondrocytes extracellular matrix degradation and inflammation in osteoarthritis via TNF/NF-κB pathway

doi: 10.1016/j.jot.2024.02.009

Figure Lengend Snippet: Sparcl1 exacerbated deteriorated subchondral bone in ACLT mice. A Representative micro-CT images of tibial plateau in coronal plane. B Representative 3D micro-CT images of subchondral trabecular bone of tibial plateau. C-J Quantitative micro-CT analysis of tibial subchondral trabecular bone: BV/TV (C), Tb. th (D), Tb.N (E), BS/TV (F), Tb. sp (G), Tb. Pf (H), BS/BV (I) and SMI (J). BV/TV, Bone Volume/Total Volume; Tb.Th, Trabecular Bone Number; Tb.N, Trabecular Bone Number; BS/TV, Bone Surface/Total Volume; Tb. Sp, Trabecular Bone Separation; Tb. Pf, Trabecular Bone pattern factor; BS/BV, Bone Surface/Bone Volume; SMI, structure model index. n = 5 in each group. Data are expressed as mean ± SD, *P < 0.05; **P < 0.01; ***P < 0.001. One-way ANOVA with post-hoc Bonferroni correction was used for comparisons.

Article Snippet: Proteins were analyzed with antibodies against matrix metalloproteinases 3 (MMP3) (1:1000, Santa Cruz Cat# sc-21732), ADAMTS5 (1:1 000, Affinity Cat# DF13268), FAK (1:1000, Cell Signaling Technology Cat# 3285), Phospho-FAK (1:1000, Cell Signaling Technology Cat# 3285), NFκB (1:1000, Cell Signaling Technology Cat# 6956), IKKα (1:1000, Cell Signaling Technology Cat# 2682), IKKβ (1:1000, Cell Signaling Technology Cat# 2370), SPARCL1 (1:1000, Proteintech Cat#13517-1-AP), β-ACTIN (1:1000, Servicebio Cat# GB11001), Goat Anti-Rabbit IgG (1:5000, Jackson Cat#111–005) and Goat Anti-mouse IgG (1:5000, Jackson Cat#115–005).

Techniques: Micro-CT

Journal: Journal of Orthopaedic Translation

Article Title: SPARCL1 promotes chondrocytes extracellular matrix degradation and inflammation in osteoarthritis via TNF/NF-κB pathway

doi: 10.1016/j.jot.2024.02.009

Figure Lengend Snippet: RNA-seq analysis of Sparcl1 treated chondrocytes. A A volcano plot illustrating differentially regulated gene expression from RNA-seq analysis between ctrl and Sparcl1 treated MCCs. Genes upregulated and downregulated are shown in red and blue, respectively. B The Gene Ontology (GO) functional clustering of genes in Sparcl1-treated MCC (top 20 most significantly affected categories are shown) C-D Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis of total genes (C) and upregulated genes (D) in Sparcl1-treated MCC transcriptome. E Pathway network illustrated the relationships of each pathway enriched in KEGG analysis.

Article Snippet: Proteins were analyzed with antibodies against matrix metalloproteinases 3 (MMP3) (1:1000, Santa Cruz Cat# sc-21732), ADAMTS5 (1:1 000, Affinity Cat# DF13268), FAK (1:1000, Cell Signaling Technology Cat# 3285), Phospho-FAK (1:1000, Cell Signaling Technology Cat# 3285), NFκB (1:1000, Cell Signaling Technology Cat# 6956), IKKα (1:1000, Cell Signaling Technology Cat# 2682), IKKβ (1:1000, Cell Signaling Technology Cat# 2370), SPARCL1 (1:1000, Proteintech Cat#13517-1-AP), β-ACTIN (1:1000, Servicebio Cat# GB11001), Goat Anti-Rabbit IgG (1:5000, Jackson Cat#111–005) and Goat Anti-mouse IgG (1:5000, Jackson Cat#115–005).

Techniques: RNA Sequencing, Gene Expression, Functional Assay

Journal: Journal of Orthopaedic Translation

Article Title: SPARCL1 promotes chondrocytes extracellular matrix degradation and inflammation in osteoarthritis via TNF/NF-κB pathway

doi: 10.1016/j.jot.2024.02.009

Figure Lengend Snippet: Inflammatory response was increased in Sparcl1 treated chondrocytes. A GSEA plots evaluating the changes of inflammatory response related genes. B STRING analysis identified protein–protein interactions in the enriched pathways after Sparcl1 stimulation. C Quantification of mRNA levels for DEGs mentioned above in chondrocytes treated with Sparcl1 protein (10, 25, 50 ng/ml). Data are expressed as mean ± SD, *P < 0.05; **P < 0.01; ***P < 0.001. One-way ANOVA with post-hoc Bonferroni correction was used for comparisons.

Article Snippet: Proteins were analyzed with antibodies against matrix metalloproteinases 3 (MMP3) (1:1000, Santa Cruz Cat# sc-21732), ADAMTS5 (1:1 000, Affinity Cat# DF13268), FAK (1:1000, Cell Signaling Technology Cat# 3285), Phospho-FAK (1:1000, Cell Signaling Technology Cat# 3285), NFκB (1:1000, Cell Signaling Technology Cat# 6956), IKKα (1:1000, Cell Signaling Technology Cat# 2682), IKKβ (1:1000, Cell Signaling Technology Cat# 2370), SPARCL1 (1:1000, Proteintech Cat#13517-1-AP), β-ACTIN (1:1000, Servicebio Cat# GB11001), Goat Anti-Rabbit IgG (1:5000, Jackson Cat#111–005) and Goat Anti-mouse IgG (1:5000, Jackson Cat#115–005).

Techniques: Protein-Protein interactions

Journal: Journal of Orthopaedic Translation

Article Title: SPARCL1 promotes chondrocytes extracellular matrix degradation and inflammation in osteoarthritis via TNF/NF-κB pathway

doi: 10.1016/j.jot.2024.02.009

Figure Lengend Snippet: Sparcl1 regulated inflammation and ECM degradation through TNF/NF-κB pathway. A GSEA plots evaluating the changes of ECM disassembly related genes. B Protein expression of FAK, pFAK, ADAMTS5 and MMP3 in chondrocytes treated with recombinant Sparcl1 protein (10, 25, 50 ng/ml) for 72 h analyzed by Western blot, and quantification of immunoblots (n = 3). C GSEA plots evaluating the changes of TNFA signaling via NF-κB related genes. D-E Protein expression of TNF, IKKα, IKKβ, and NF-κB in chondrocytes treated with recombinant Sparcl1 protein (10, 25, 50 ng/ml) for 72 h analyzed by Western blot, and quantification of immunoblots (n = 3). Data are expressed as mean ± SD, *P < 0.05; **P < 0.01; ***P < 0.001. One-way ANOVA with post-hoc Bonferroni correction was used for comparisons.

Article Snippet: Proteins were analyzed with antibodies against matrix metalloproteinases 3 (MMP3) (1:1000, Santa Cruz Cat# sc-21732), ADAMTS5 (1:1 000, Affinity Cat# DF13268), FAK (1:1000, Cell Signaling Technology Cat# 3285), Phospho-FAK (1:1000, Cell Signaling Technology Cat# 3285), NFκB (1:1000, Cell Signaling Technology Cat# 6956), IKKα (1:1000, Cell Signaling Technology Cat# 2682), IKKβ (1:1000, Cell Signaling Technology Cat# 2370), SPARCL1 (1:1000, Proteintech Cat#13517-1-AP), β-ACTIN (1:1000, Servicebio Cat# GB11001), Goat Anti-Rabbit IgG (1:5000, Jackson Cat#111–005) and Goat Anti-mouse IgG (1:5000, Jackson Cat#115–005).

Techniques: Expressing, Recombinant, Western Blot

Journal: Journal of Orthopaedic Translation

Article Title: SPARCL1 promotes chondrocytes extracellular matrix degradation and inflammation in osteoarthritis via TNF/NF-κB pathway

doi: 10.1016/j.jot.2024.02.009

Figure Lengend Snippet: BAY 11 – 7082 could reverse Sparcl1 induced ECM degradation and related gene expression. A Alcian blue staining of ATDC5 with micromass culture in chondrogenesis media with or without recombinant Sparcl1 protein and BAY 11–7082 (2.5 μM). Representative image of staining at day 8 is shown. B Quantification of mRNA levels for Col2a1, Acan, Sox9, Mmp3, Mmp13, Adamts5 in MCC treated with recombinant Sparcl1 protein (50 ng/ml) and BAY 11–7082 (2.5 μM) for 48 h. C Immunoblot analysis of MCC, immunoprecipitated with anti-TNFR and IgG and analyzed by immunoblot with anti-SPARCL1.

Article Snippet: Proteins were analyzed with antibodies against matrix metalloproteinases 3 (MMP3) (1:1000, Santa Cruz Cat# sc-21732), ADAMTS5 (1:1 000, Affinity Cat# DF13268), FAK (1:1000, Cell Signaling Technology Cat# 3285), Phospho-FAK (1:1000, Cell Signaling Technology Cat# 3285), NFκB (1:1000, Cell Signaling Technology Cat# 6956), IKKα (1:1000, Cell Signaling Technology Cat# 2682), IKKβ (1:1000, Cell Signaling Technology Cat# 2370), SPARCL1 (1:1000, Proteintech Cat#13517-1-AP), β-ACTIN (1:1000, Servicebio Cat# GB11001), Goat Anti-Rabbit IgG (1:5000, Jackson Cat#111–005) and Goat Anti-mouse IgG (1:5000, Jackson Cat#115–005).

Techniques: Gene Expression, Staining, Recombinant, Western Blot, Immunoprecipitation

Journal: Journal of Orthopaedic Translation

Article Title: SPARCL1 promotes chondrocytes extracellular matrix degradation and inflammation in osteoarthritis via TNF/NF-κB pathway

doi: 10.1016/j.jot.2024.02.009

Figure Lengend Snippet: Mechanism of Sparcl1 in exacerbate osteoarthritis. Diagram illustrating the mechanism how Sparcl1 influences OA pathogenesis through the TNF/NFκB signaling pathway.

Article Snippet: Proteins were analyzed with antibodies against matrix metalloproteinases 3 (MMP3) (1:1000, Santa Cruz Cat# sc-21732), ADAMTS5 (1:1 000, Affinity Cat# DF13268), FAK (1:1000, Cell Signaling Technology Cat# 3285), Phospho-FAK (1:1000, Cell Signaling Technology Cat# 3285), NFκB (1:1000, Cell Signaling Technology Cat# 6956), IKKα (1:1000, Cell Signaling Technology Cat# 2682), IKKβ (1:1000, Cell Signaling Technology Cat# 2370), SPARCL1 (1:1000, Proteintech Cat#13517-1-AP), β-ACTIN (1:1000, Servicebio Cat# GB11001), Goat Anti-Rabbit IgG (1:5000, Jackson Cat#111–005) and Goat Anti-mouse IgG (1:5000, Jackson Cat#115–005).

Techniques: