Journal: Neuropsychiatric Disease and Treatment

Article Title: Neuroprotective Function of TNFAIP3 Interacting Protein 2 Against Oxygen and Glucose Deprivation/Reoxygenation-Induced Injury in Hippocampal Neuronal HT22 Cells Through Regulation of the TLR4/MyD88/NF-κB Pathway

doi: 10.2147/ndt.s308360

Figure Lengend Snippet: Figure 1 Expression of TNIP2 in HT-22 cells subjected to OGD/R. (A) qRT-PCR analysis of TNIP2 expression showed the reduced level of TNIP2 mRNA in HT22 cells exposed to OGD/R (n = 3). (B) Western blot analysis of TNIP2 expression showed the reduced level of TNIP2 protein in HT22 cells exposed to OGD/R (n = 3). **P <0.01 and ***P <0.001, compared with the control group.

Article Snippet: Subsequently, the members were blocked for 1 h with 5% non-fat milk, and then probed at 4°C overnight with the special primary antibodies against TNIP2 (Novus, Shanghai, China), toll-like receptor 4 https://doi.org/10.2147/NDT.S308360 DovePress Neuropsychiatric Disease and Treatment 2021:17 2220 Powered by TCPDF (www.tcpdf.org) (TLR4; Novus), myeloid differentiation factor 88 (MyD88; Novus), NF-кB (Novus), and β-actin (Novus).

Techniques: Expressing, Quantitative RT-PCR, Western Blot, Control

Journal: Neuropsychiatric Disease and Treatment

Article Title: Neuroprotective Function of TNFAIP3 Interacting Protein 2 Against Oxygen and Glucose Deprivation/Reoxygenation-Induced Injury in Hippocampal Neuronal HT22 Cells Through Regulation of the TLR4/MyD88/NF-κB Pathway

doi: 10.2147/ndt.s308360

Figure Lengend Snippet: Figure 2 TNIP2 overexpression attenuates ODG/R-induced apoptosis in HT22 cells. (A) LV-Scramble and LV-TNIP2 were transfected into HT22 cells for 48 h and the transfection efficiency was confirmed using Western blot (n = 3). HT22 cells were transfected with LV-TNIP2 or LV-Scramble, followed by exposure to OGD/R. (B and C) The expression levels of TNIP2 protein and mRNA were detected by Western blot and qRT-PCR, respectively. (D) The viability of HT22 cells was examined using CCK-8 assay (n = 3). (E) The viability of HT22 cells was determined by measurement of LDH release (n = 3). (F) Representative images of TUNEL staining were shown and the bar graph represents the quantification of TUNEL positive cells (n = 3). (G) Caspase-3 activity assay kit was applied to examine the activity of caspase-3 (n = 3). **P <0.01 and ***P <0.001, compared with the LV-Scramble or control group. #P <0.05, ##P <0.01, and ###P <0.001, compared with the LV-Scramble + OGD/R group.

Article Snippet: Subsequently, the members were blocked for 1 h with 5% non-fat milk, and then probed at 4°C overnight with the special primary antibodies against TNIP2 (Novus, Shanghai, China), toll-like receptor 4 https://doi.org/10.2147/NDT.S308360 DovePress Neuropsychiatric Disease and Treatment 2021:17 2220 Powered by TCPDF (www.tcpdf.org) (TLR4; Novus), myeloid differentiation factor 88 (MyD88; Novus), NF-кB (Novus), and β-actin (Novus).

Techniques: Over Expression, Transfection, Western Blot, Expressing, Quantitative RT-PCR, CCK-8 Assay, TUNEL Assay, Staining, Caspase-3 Activity Assay, Activity Assay, Control

Journal: Neuropsychiatric Disease and Treatment

Article Title: Neuroprotective Function of TNFAIP3 Interacting Protein 2 Against Oxygen and Glucose Deprivation/Reoxygenation-Induced Injury in Hippocampal Neuronal HT22 Cells Through Regulation of the TLR4/MyD88/NF-κB Pathway

doi: 10.2147/ndt.s308360

Figure Lengend Snippet: Figure 3 TNIP2 overexpression attenuates ODG/R-induced inflammation in HT22 cells. ELISA assays were conducted to evaluate the levels of TNF-α (A), IL-1β (B), ICAM- 1 (C), and IL-10 (D) in HT22 cells transfected with LV-TNIP2 or LV-Scramble after OGD/R (n = 3). (E) The activity of NF-кB was determined in HT22 cells transfected with LV-TNIP2 or LV-Scramble after OGD/R (n = 3). **P <0.01 and ***P <0.001, compared with the control group. #P <0.05 and ##P <0.01 compared with the LV-Scramble + OGD/R group.

Article Snippet: Subsequently, the members were blocked for 1 h with 5% non-fat milk, and then probed at 4°C overnight with the special primary antibodies against TNIP2 (Novus, Shanghai, China), toll-like receptor 4 https://doi.org/10.2147/NDT.S308360 DovePress Neuropsychiatric Disease and Treatment 2021:17 2220 Powered by TCPDF (www.tcpdf.org) (TLR4; Novus), myeloid differentiation factor 88 (MyD88; Novus), NF-кB (Novus), and β-actin (Novus).

Techniques: Over Expression, Enzyme-linked Immunosorbent Assay, Transfection, Activity Assay, Control

Journal: Neuropsychiatric Disease and Treatment

Article Title: Neuroprotective Function of TNFAIP3 Interacting Protein 2 Against Oxygen and Glucose Deprivation/Reoxygenation-Induced Injury in Hippocampal Neuronal HT22 Cells Through Regulation of the TLR4/MyD88/NF-κB Pathway

doi: 10.2147/ndt.s308360

Figure Lengend Snippet: Figure 4 The TLR4/MyD88/NF-κB pathway is involved in the protective effects of TNIP2 on OGD/R-induced neuronal damage. HT22 cells were transfected with LV-TNIP2 or LV-Scramble, followed by exposure to OGD/R. (A) Representative and (B) quantitative Western blot analysis of TLR4, MyD88, NF-κB-cytoplasm, and NF-κB-nucleus expression in HT22 cells transfected with LV-TNIP2 or LV-Scramble following OGD/R (n = 3). ***P <0.001, compared with the control group. ##P <0.01, compared with the LV-Scramble + OGD/R group.

Article Snippet: Subsequently, the members were blocked for 1 h with 5% non-fat milk, and then probed at 4°C overnight with the special primary antibodies against TNIP2 (Novus, Shanghai, China), toll-like receptor 4 https://doi.org/10.2147/NDT.S308360 DovePress Neuropsychiatric Disease and Treatment 2021:17 2220 Powered by TCPDF (www.tcpdf.org) (TLR4; Novus), myeloid differentiation factor 88 (MyD88; Novus), NF-кB (Novus), and β-actin (Novus).

Techniques: Transfection, Western Blot, Expressing, Control

Journal: Journal of Virology

Article Title: Foot-and-Mouth Disease Virus Structural Protein VP1 Destroys the Stability of the TPL2 Trimer by Degradation of TPL2 To Evade Host Antiviral Immunity

doi: 10.1128/JVI.02149-20

Figure Lengend Snippet: TPL2, ABIN2, and P105 levels were decreased after FMDV infection in vivo and in vitro . (A) PK-15 cells were infected with equal amounts of FMDV (MOI of 1) for 0, 4, 8, or 12 h. The transcriptional levels of FMDV and TPL2 were measured by RT-qPCR. (B) PK-15 cells were infected with equal amounts of FMDV (MOI of 1) for 0, 2, 4, 8, 12, 16, and 20 h. The expression levels of endogenous TPL2 and viral proteins were detected by Western blotting with anti-TPL2 and anti-FMDV antibodies. (C) The mRNA expression levels of p105 and ABIN2 were measured by RT-qPCR for the samples from panel A. (D) The expression levels of endogenous p105, ABIN2, and viral proteins were detected by Western blotting for the samples from panel B with anti-p105, anti-ABIN2, and anti-FMDV antibodies. (E) PK-15 cells with or without actinomycin D (10 μg/ml) treatment were infected with equal amounts of FMDV (MOI of 1) for 0, 4, 8, 12, and 16 h. The expression levels of endogenous TPL2 and viral proteins were detected by Western blotting. (F and G) Three-day-old wild-type mice ( n = 4 in each group) were divided into two groups. Two groups were subcutaneously injected with equal amounts of FMDV (7 × 10 4 PFU) and PBS and then euthanized 72 h after FMDV infection. All Western blotting samples from suckling mice without FMDV infection were mixed, and all Western blotting samples of suckling mice infected with FMDV were not mixed. (F) Expression levels of TPL2, p105, and ABIN2 proteins in mice were detected by Western blotting with anti-TPL2, anti-p105, and anti-ABIN2 antibodies. Relative fold change in abundance of TPL2, p105, and ABIN2 protein was determined by densitometric analysis. WT-mice1, all combinations of noninfected FMDV suckling mice; WT-mice2, WT-mice3, and WT-mice4, three suckling mice infected with FMDV. (G) RT-qPCR samples of uninfected suckling mice and FMDV-infected suckling mice were not mixed. The transcriptional levels of TPL2, p105, and ABIN2 in mice were measured by RT-qPCR.

Article Snippet: The commercial antibodies used in this study were mouse anti-Flag monoclonal antibody, mouse anti-Myc monoclonal antibody, mouse anti-HA monoclonal antibody (all from Sigma-Aldrich), mouse anti-V5 monoclonal antibody (Proteintech), mouse anti-TPL2 monoclonal antibody (Santa Cruz Biotechnology), mouse anti-eIF4G monoclonal antibody (Santa Cruz Biotechnology), rabbit anti-p105 polyclonal antibody (Cell Signaling Technology), rabbit anti-ABIN2 polyclonal antibody (Proteintech), anti-β-actin monoclonal antibody (Thermo Fisher Scientific), goat anti-mouse IgG antibody (Proteintech), and goat anti-rabbit IgG antibody (Proteintech).

Techniques: Infection, In Vivo, In Vitro, Quantitative RT-PCR, Expressing, Western Blot, Injection

Journal: Journal of Virology

Article Title: Foot-and-Mouth Disease Virus Structural Protein VP1 Destroys the Stability of the TPL2 Trimer by Degradation of TPL2 To Evade Host Antiviral Immunity

doi: 10.1128/JVI.02149-20

Figure Lengend Snippet: FMDV VP1 protein was responsible for reducing the expression of TPL2, p105, and ABIN2 at protein level but not transcription level. (A) HEK293T cells were cotransfected with Myc-TPL2 (1 μg), along with increasing amounts of Flag-L-, Flag-VP1-, or Flag-VP2-expressing plasmid (0, 1, 3, and 5 μg), and the empty Flag vector (5, 4, 2, and 0 μg) was used in the transfection process. At 24 hpt, the expression levels of Myc-TPL2 and Flag-tagged viral proteins (L, VP1, and VP2) were analyzed by Western blotting with anti-Myc and anti-Flag antibodies. (B) PK-15 cells were transfected with increasing amounts of Flag-L-, Flag-VP1-, or Flag-VP2-expressing plasmid (0, 1, 3, and 5 μg), and the empty Flag vector (5, 4, 2, and 0 μg) was used in the transfection process. At 24 hpt, the expression levels of endogenous TPL2 and Flag-tagged viral proteins were analyzed by Western blotting with anti-TPL2 and anti-Flag antibodies. (C) HEK293T cells were cotransfected with HA-p105 or Flag-ABIN2 (1 μg), along with increasing amounts of Flag-VP1- or HA-VP1-expressing plasmid (0, 1, 3, and 5 μg), and the empty Flag vector or empty HA vector (5, 4, 2, and 0 μg) was used in the transfection process. At 24 hpt, the expression levels of HA-p105 and Flag-ABIN2 proteins were analyzed by Western blotting with anti-HA and anti-Flag antibodies. (D and E) PK-15 cells were transfected with increasing amounts of Flag-VP1-expressing plasmid (0, 1, 3, or 5 μg) for 24 h. Empty Flag vector (5, 4, 2, or 0 μg) was used in the transfection process. (D) The expression levels of endogenous p105 and ABIN2 proteins were detected by Western blotting with anti-p105 and anti-ABIN2 antibodies. (E) The mRNA expression levels of endogenous TPL2, p105, and ABIN2 were measured by RT-qPCR. (F) PK-15 cells were transfected with FMDV L-expressing plasmid (3 μg), and the cells were collected at 0, 16, 32, and 48 hpt. The protein levels of eIF4G and Flag-L were determined by Western blotting. (G and H) PK-15 cells with or without infection with FMDV type A strain-WT, FMDV type O strain-WT, or FMDV type O strain-ΔL (MOI of 1) for 12 h. (G) The expression levels of endogenous TPL2 were detected by Western blotting. (H) The expression levels of TPL2 mRNA were measured by RT-qPCR.

Article Snippet: The commercial antibodies used in this study were mouse anti-Flag monoclonal antibody, mouse anti-Myc monoclonal antibody, mouse anti-HA monoclonal antibody (all from Sigma-Aldrich), mouse anti-V5 monoclonal antibody (Proteintech), mouse anti-TPL2 monoclonal antibody (Santa Cruz Biotechnology), mouse anti-eIF4G monoclonal antibody (Santa Cruz Biotechnology), rabbit anti-p105 polyclonal antibody (Cell Signaling Technology), rabbit anti-ABIN2 polyclonal antibody (Proteintech), anti-β-actin monoclonal antibody (Thermo Fisher Scientific), goat anti-mouse IgG antibody (Proteintech), and goat anti-rabbit IgG antibody (Proteintech).

Techniques: Expressing, Plasmid Preparation, Transfection, Western Blot, Quantitative RT-PCR, Infection

Journal: Journal of Virology

Article Title: Foot-and-Mouth Disease Virus Structural Protein VP1 Destroys the Stability of the TPL2 Trimer by Degradation of TPL2 To Evade Host Antiviral Immunity

doi: 10.1128/JVI.02149-20

Figure Lengend Snippet: VP1 interacted with host TPL2 but not with p105 and ABIN2. (A) HEK293T cells were cotransfected with 8 μg Myc-TPL2- or HA-p105- and Flag-VP1-expressing plasmid. The cells were lysed at 24 hpt, and the lysates were immunoprecipitated with mouse IgG antibody or mouse anti-Flag antibody and subjected to Western blotting. Lysate and immunoprecipitation (IP) antibody-antigen complexes were analyzed by immunoblotting (IB) using anti-HA, anti-Myc, anti-Flag, and anti-β-actin antibodies. (B) The method is the same as that for panel A. The lysates were immunoprecipitated with mouse IgG antibody or mouse anti-HA antibody and subjected to Western blotting. (C and D) PK-15 cells were transfected with Flag-VP1-expressing plasmid (8 μg) or empty Flag vector (Vec; 8 μg) for 24 h. The cells were then lysed and immunoprecipitated with normal IgG antibody, anti-Flag antibody (C), or anti-TPL2 antibody (D). The lysate and IP antibody-antigen complexes were analyzed by IB using anti-Flag, anti-TPL2, or anti-β-actin antibodies. (E and F) PK-15 cells were mock infected or infected with FMDV (MOI of 1) for 6 or 12 h. MG132 (20 μM) was added to prevent the degradation of endogenous TPL2. The cells were then lysed and immunoprecipitated with normal IgG antibody, anti-VP1 antibody (E), or anti-TPL2 antibody (F). The lysate and IP antibody-antigen complexes were analyzed by IB using anti-TPL2, anti-VP1, or anti-β-actin antibodies. (G) Schematic representation showing a series of Myc-tagged truncated TPL2 mutants. (H) Schematic representation showing a series of Flag-tagged truncated VP1 mutants. (I and J) HEK293T cells were cotransfected with HA-VP1 (7 μg) and a series of Myc-tagged TPL2 truncated mutants expressing plasmids (7 μg). The cells were lysed at 24 hpt, and the lysates were immunoprecipitated with mouse IgG antibody, mouse anti-HA antibody (I), or mouse anti-Myc antibody (J) and subjected to Western blotting. (K and L) HEK293T cells were cotransfected with Myc-TPL2 (7 μg) and a series of Flag-tagged VP1 truncated mutants expressing plasmids (7 μg). The cells were lysed at 24 hpt, and the lysates were immunoprecipitated with mouse IgG antibody, mouse anti-Myc antibody (K), or mouse anti-Flag antibody (L) and subjected to Western blotting.

Article Snippet: The commercial antibodies used in this study were mouse anti-Flag monoclonal antibody, mouse anti-Myc monoclonal antibody, mouse anti-HA monoclonal antibody (all from Sigma-Aldrich), mouse anti-V5 monoclonal antibody (Proteintech), mouse anti-TPL2 monoclonal antibody (Santa Cruz Biotechnology), mouse anti-eIF4G monoclonal antibody (Santa Cruz Biotechnology), rabbit anti-p105 polyclonal antibody (Cell Signaling Technology), rabbit anti-ABIN2 polyclonal antibody (Proteintech), anti-β-actin monoclonal antibody (Thermo Fisher Scientific), goat anti-mouse IgG antibody (Proteintech), and goat anti-rabbit IgG antibody (Proteintech).

Techniques: Expressing, Plasmid Preparation, Immunoprecipitation, Western Blot, Transfection, Infection

Journal: Journal of Virology

Article Title: Foot-and-Mouth Disease Virus Structural Protein VP1 Destroys the Stability of the TPL2 Trimer by Degradation of TPL2 To Evade Host Antiviral Immunity

doi: 10.1128/JVI.02149-20

Figure Lengend Snippet: Degradation pathways of TPL2, ABIN2, and p105 induced by VP1 in HEK293T and PK-15 cells. (A) PK-15 cells treated with MG132 (20 μM), CQ (100 μM), NH 4 Cl (20 μM), or Z-VAD-FMK (50 μM) for 1 h and then with or without FMDV infection (MOI of 1) for 12 h. The expression levels of endogenous TPL2 and viral proteins were detected by Western blotting. DMSO, dimethyl sulfoxide. (B) HEK293T cells were cotransfected with Flag-VP1-expressing plasmid (2 μg) or empty Flag vector (2 μg) and Myc-TPL2-expressing plasmid (1 μg) and empty Myc vector (1 μg) and maintained in the presence or absence of MG132 (2 or 20 μM), CQ (50 or 100 μM), NH 4 Cl (10 or 20 μM), or Z-VAD-FMK (10 or 50 μM) for 36 h. The expression levels of Myc-TPL2 and Flag-VP1 proteins were detected by Western blotting. (C) PK-15 cells were transfected with HA-VP1-expressing plasmid (2 μg) or empty HA vector (2 μg) and maintained in the presence or absence of MG132 (20 μM), CQ (100 μM), NH 4 Cl (20 μM), or Z-VAD-FMK (50 μM) for 36 h. The expression levels of endogenous TPL2 and HA-VP1 proteins were detected by Western blotting. (D) HEK293T cells were cotransfected with HA-VP1 (5 μg) or empty HA vector (5 μg) and Myc-tagged TPL2 truncated mutant-expressing plasmids (2 μg). At 24 hpt, the cells were collected for Western blotting. (E) HEK293T cells were cotransfected with HA-VP1 (1 μg), along with increasing amounts of Myc-TPL2-expressing plasmid (0, 1, 2, and 3 μg), and the empty Myc vector (3, 2, 1, or 0 μg) was used in the transfection process. At 24 hpt, the cells were collected for Western blotting. (F) PK-15-TPL2 or PK-15-TPL2 −/− cells were transfected with HA-VP1-expressing plasmid (2 μg) or empty HA vector (2 μg). At 24 hpt, the cells were collected for Western blotting. (G and H) Similar transfection and immunoblotting analyses in HEK293T and PK-15 cells were performed as described for and . (I) HEK293T cells were cotransfected with Flag-P1 (3 μg) or empty Flag vector (3 μg) and Myc-TPL2-expressing plasmid (1 μg) and maintained in the presence or absence of MG132 (20 μM), CQ (100 μM), NH 4 Cl (20 μM), and Z-VAD-FMK (50 μM) for 36 h. The expression levels of Myc-TPL2 and Flag-P1 proteins were detected by Western blotting. PK-15 cells were transfected with Flag-P1-expressing plasmid (3 μg) or empty Flag vector (3 μg) and maintained in the presence or absence of MG132 (20 μM), CQ (100 μM), NH 4 Cl (20 μM), and Z-VAD-FMK (50 μM) for 36 h. The expression levels of endogenous TPL2 and Flag-P1 proteins were detected by Western blotting. (J) HEK293T cells were cotransfected with Flag-Ub-expressing (4 μg), Myc-TPL2-expressing (4 μg), HA-VP1-expressing (6 μg) plasmid, and empty HA vector (6 μg) for 20 h. The cells were then lysed and immunoprecipitated with mouse IgG antibody or mouse anti-Myc antibody and subjected to Western blotting. (K) HEK293T cells were cotransfected with HA-Ub, HA-K48-Ub, or HA-K63-Ub (4 μg), Myc-TPL2 (4 μg), Flag-VP1-expressing plasmid (6 μg), and empty Flag vector (6 μg) for 20 h. The cells were then lysed and immunoprecipitated with mouse anti-Myc antibody and subjected to Western blotting. (L) Degradation pathway of p105 induced by VP1 in HEK293T and PK-15 cells. The method is the same as that for panel I. (M) Degradation pathway of ABIN2 induced by VP1 in HEK293T and PK-15 cells. The method is the same as that for panel I.

Article Snippet: The commercial antibodies used in this study were mouse anti-Flag monoclonal antibody, mouse anti-Myc monoclonal antibody, mouse anti-HA monoclonal antibody (all from Sigma-Aldrich), mouse anti-V5 monoclonal antibody (Proteintech), mouse anti-TPL2 monoclonal antibody (Santa Cruz Biotechnology), mouse anti-eIF4G monoclonal antibody (Santa Cruz Biotechnology), rabbit anti-p105 polyclonal antibody (Cell Signaling Technology), rabbit anti-ABIN2 polyclonal antibody (Proteintech), anti-β-actin monoclonal antibody (Thermo Fisher Scientific), goat anti-mouse IgG antibody (Proteintech), and goat anti-rabbit IgG antibody (Proteintech).

Techniques: Infection, Expressing, Western Blot, Plasmid Preparation, Transfection, Mutagenesis, Immunoprecipitation

Journal: Journal of Virology

Article Title: Foot-and-Mouth Disease Virus Structural Protein VP1 Destroys the Stability of the TPL2 Trimer by Degradation of TPL2 To Evade Host Antiviral Immunity

doi: 10.1128/JVI.02149-20

Figure Lengend Snippet: FMDV and FMDV VP1 destroy the stability of the TPL2-p105-ABIN2 trimer complex. (A) PK-15 cells were cotransfected in pairs with plasmid expressing Myc-TPL2 (5 μg), HA-p105 (5 μg), or Flag-ABIN (5 μg) for 24 h, with or without FMDV infection (MOI of 1) for 12 h. The cells were lysed, and the lysates were immunoprecipitated with anti-Myc antibody, mouse anti-Flag antibody, anti-HA antibody, and IgG antibody and subjected to Western blotting. (B) PK-15 cells with or without FMDV infection (MOI of 1) for 12 h. The cells were lysed and immunoprecipitated with mouse anti-TPL2 antibody. The lysate and IP antibody-antigen complexes were analyzed by IB using anti-TPL2, anti-p105, anti-ABIN2, anti-FMDV, and anti-β-actin antibodies to measure the expression of endogenous TPL2, p105, and ABIN2 separately. (C) HEK293T cells were cotransfected in pairs with plasmid expressing Myc-TPL2 (5 μg), HA-p105 (5 μg), and V5-ABIN2 (5 μg). Flag-VP1-expressing plasmid (8 μg) and empty Flag vector (8 μg) were transfected at the same time. At 24 hpt, the cells were lysed and the lysates were immunoprecipitated with anti-Myc antibody, anti-V5 antibody, anti-HA antibody, and mouse normal IgG antibody. (D) PK-15 cells were transfected with Flag-VP1-expressing plasmid (8 μg) and empty Flag vector (8 μg) for 24 h. The cells were lysed and immunoprecipitated with anti-TPL2 antibody and subjected to Western blotting. The lysate and IP antibody-antigen complexes were analyzed by IB to measure the expression of endogenous TPL2, p105, and ABIN2 separately.

Article Snippet: The commercial antibodies used in this study were mouse anti-Flag monoclonal antibody, mouse anti-Myc monoclonal antibody, mouse anti-HA monoclonal antibody (all from Sigma-Aldrich), mouse anti-V5 monoclonal antibody (Proteintech), mouse anti-TPL2 monoclonal antibody (Santa Cruz Biotechnology), mouse anti-eIF4G monoclonal antibody (Santa Cruz Biotechnology), rabbit anti-p105 polyclonal antibody (Cell Signaling Technology), rabbit anti-ABIN2 polyclonal antibody (Proteintech), anti-β-actin monoclonal antibody (Thermo Fisher Scientific), goat anti-mouse IgG antibody (Proteintech), and goat anti-rabbit IgG antibody (Proteintech).

Techniques: Plasmid Preparation, Expressing, Infection, Immunoprecipitation, Western Blot, Transfection