Skin tissue sections were deparaffinized, rehydrated and blocked with 3% H2O2 for 15 min. The sections were incubated with primary antibodies such as anti-HGF (1:1200, DF6326, Affinty Biosciences, China), anti-Nrf2 (1:3000, 16 396–1-AP, Proteintech, China), anti-Bcl-2 (1:100, A19693, Abclonal, China), anti-Bax (1:5000, A19684, Abclonal, China) or anti-vascular endothelial growth factor (VEGF; 1:7000, 19 003–1-AP, Proteintech, China) at 4°C overnight. After thorough washing, sections were incubated with horseradish peroxidase-conjugated secondary antibody (1:5000, Proteintech, China) for 30 min at room temperature, then stained with diaminobenzidine solution. Sections were visualized under a microscope, and representative sections were scanned and analysed using a biopsy scanner (NDP, Hamamatsu, Japan). ImageJ software quantified the percentage contribution of the positive with four random images were taken from each group.
A19684
Manufactured by ABclonal
Sourced in China
A19684 is a centrifuge machine used for separating different components of a liquid mixture based on their density and size. It is designed for routine laboratory applications that require efficient and reliable separation of samples.
Automatically generated - may contain errors
Lab products found in correlation
A19693, by ABclonal (3 mentions)
Bca protein assay kit, by Thermo Fisher Scientific (2 mentions)
Horseradish peroxidase conjugated secondary antibody, by Proteintech (1 mentions)
Ac026, by ABclonal (1 mentions)
Sa0001 2, by Proteintech (1 mentions)
A2547, by ABclonal (1 mentions)
Sc 47778, by Santa Cruz Biotechnology (1 mentions)
A0216, by Beyotime (1 mentions)
Ap0707, by ABclonal (1 mentions)
A02177 4, by Boster Bio (1 mentions)
A19714, by ABclonal (1 mentions)
Ap0123, by ABclonal (1 mentions)
Af7022, by Affinity Biosciences (1 mentions)
β actin, by Santa Cruz Biotechnology (1 mentions)
Bca protein assay kit, by Beyotime (1 mentions)
A0208, by Beyotime (1 mentions)
Ab194583, by Abcam (1 mentions)
Bs 23003r, by Bioss Antibodies (1 mentions)
As1107, by Aspen (1 mentions)
Ab32101, by Abcam (1 mentions)
6 protocols using a19684
1
Immunohistochemical Analysis of Skin Tissue
2
Western Blot Analysis of Protein Expression
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HaCaT cells were harvested with protein lysis buffer. And the protein concentrations were assessed using bicinchoninic acid (BCA) Protein Assay kit (Thermo Fisher Scientific, Waltham, MA). Protein samples were separated by 10% sodium dodecyl sulfate-polyacrylamide gel electrophoresis at 100 V for 2 h, and then transferred to nitrocellulose membranes. After blocking with 5% dried nonfat milk for 1 h at room temperature, the membranes were incubated with primary antibodies, such as anti-Nrf2 (1:1000, 16396-1-AP, Proteintech, China), anti-Bcl-2 (1:1000, A19693, Abclonal, China), anti-Bax (1:1000, A19684, Abclonal, China) or anti-β-actin (1:10 000, AC026, Abclonal, China), respectively, overnight at 4°C. Following three rinses, blots were incubated with horseradish peroxidase-conjugated secondary antibodies (1:10 000, SA0001-2, Proteintech, China) for 1 h at room temperature. Then the chemiluminescence signal in the blots was detected with Hypersensitive Chemiluminescence Substrate kit (17046, Zenbio, China) using the ePhoto developer (GenScript, China). The relative expression of the target protein was assessed using the internal reference β-actin as the standard. The intensity was quantified by ImageJ software (National Institutes of Health, Germany).
3
Western Blot Analysis of Apoptosis and NF-κB Signaling
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Samples were lysed in cell lysis buffer for western and IP (Beyotime). Proteins were quantified by BCA protein assay kit (Beyotime). After complete separation by electrophoresis, the proteins were transferred to polyvinylidene fluoride (PVDF) membranes (Millipore, Billerica, MA, USA). Then these membranes were blocked in 5% skimmed milk (Inner Mongolia Yili Industrial Group Co., Ltd., Inner Mongolia, China) for 1 h and incubated with primary antibodies. Then these membranes were incubated with HRP-labeled goat anti-rabbit IgG (H+L) (1: 5,000, A0208, Beyotime) or horseradish peroxidase (HRP)-labeled goat anti-mouse IgG (H+L) (1: 5,000, A0216, Beyotime) at room temperature at 37°C for 45 min. Protein bands were visualized with Gel-Pro-Analyzer software (Beijing Liuyi Biotechnology Co., Ltd., Beijing, China).
The primary antibodies used were shown as following: HDAC9 antibody (1: 500, A02177-4, BOSTER, Wuhan, China), cleaved caspase-3 antibody (1:1,000, AF7022, Affinity, Jiangsu, China), Bax antibody (1:1,000, A19684, ABclonal, Wuhan, China), B-cell lymphoma/leukemia-2 (Bcl-2) antibody (1:1,000, A19693, ABclonal), IκBα antibody (1:1,000, A19714, ABclonal), p-IκBα antibody (1:1,000, AP0707, ABclonal), p-p65 antibody (1:1,000, AP0123, ABclonal), p65 antibody (1:1,000, A2547, ABclonal), β-actin (1:1,000, sc-47778, Santa Cruz, Santa Cruz, CA, USA).
The primary antibodies used were shown as following: HDAC9 antibody (1: 500, A02177-4, BOSTER, Wuhan, China), cleaved caspase-3 antibody (1:1,000, AF7022, Affinity, Jiangsu, China), Bax antibody (1:1,000, A19684, ABclonal, Wuhan, China), B-cell lymphoma/leukemia-2 (Bcl-2) antibody (1:1,000, A19693, ABclonal), IκBα antibody (1:1,000, A19714, ABclonal), p-IκBα antibody (1:1,000, AP0707, ABclonal), p-p65 antibody (1:1,000, AP0123, ABclonal), p65 antibody (1:1,000, A2547, ABclonal), β-actin (1:1,000, sc-47778, Santa Cruz, Santa Cruz, CA, USA).
4
Immunohistochemical analysis of JAK2/STAT3
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The paraffin retinal sections were processed according to IHC protocol and previous study (17 (link)). Photos were taken under an optical microscope (OLYMPUS, CX-21, Japan). Primary antibodies were as follows: JAK2 (bs-23003r, Bioss), STAT3 (60199-1-lg, Proteintech), p-JAK2 (ab32101, Abcam), p-STAT3 (sc-8059, Santa), Bcl-2 (ab194583, Abcam), and Bax (A19684, ABclonal). HRP-conjugated secondary antibodies were AS-1107 and AS-11069 (Aspen).
5
Protein Expression Analysis of Neuroinflammation
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The BV2 or SH-SY5Y cells or midbrain tissue lysates were subjected to Western blotting to detect NFATc2, STAT1, p-STAT1, IRF9, BAX, BCL-2, p65, p-p65, IκBα, p-IκBα and GAPDH. The following antibodies were used for protein detection by western blotting. NFATc2 (22023–1-AP, 1:1000, Proteintech), STAT1 (66545–1-lg, 1:2000, Proteintech), p-STAT1 (ab109461, 1:1000, Abcam), IRF9 (14167–1-AP, 1:1000, proteintech), BAX (A19684, 1:1000, ABclonal), BCL-2 (26593–1-AP, 1:1000, Proteintech), NF-kB p65 (AF5006, 1:1000, Affinity), Phospho-NF-kB p65(Ser536) (AF2006, 1:1000, Affinity), IKB alpha (AF5002, 1:1000, Affinity), Phospho-IKB alpha (Ser32/Ser36) (AF2002, 1:1000, Affinity), and GAPDH (#5174, 1:1000, Cell Signaling Technology).
6
Western Blot Analysis of Apoptosis Markers
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Total proteins were extracted using RIPA buffer (Solarbio). Sample proteins were separated by SDS gels and transferred to PVDF membranes (Millipore, USA). The membranes were probed with primary antibodies by LAMP3 antibody (1:500, 12632-1-AP, Proteintech, China), Bcl2 antibody (1:500, A19693, ABclonal, China), Bcl-xL antibody (1:1000, A19703, ABclonal), Bax antibody (1:500, A19684, ABclonal). GAPDH (1:10000, 60004-1-Ig, Proteintech) was used as an internal control. Chemiluminescence detection was performed using goat anti-rabbit IgG/HRP antibody (1:3000, SE134, Solarbio Science & Technology, Co., Ltd.) or goat anti-mouse IgG/HRP antibody (1:3000, SE131, Solarbio). The immunoreactive protein bands were visualized using ECL reagent (Solarbio).
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