Invitrogen

PINK1 Polyclonal Antibody

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问题与答案

问题与答案

PINK1 Antibody in Immunocytochemistry (ICC/IF)

PINK1 Antibody in Immunocytochemistry (ICC/IF)

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PINK1 Antibody (PA5-86941) in ICC/IF

Immunofluorescence analysis of PINK1 was performed using 70% confluent log phase SH-SY5Y treated with 25 mM Hydrogen Peroxide for 24 hours. The cells were fixed with 4% paraformaldehyde for 10 minutes, permeabilized with 0.1% Triton™ X-100 for 15 minutes, and blocked with 2% BSA for 45 minutes at room temperature. The cells were labeled with PINK1 Polyclonal Antibody (Product # PA5-86941) at 1:200 dilution in 0.1% BSA, incubated at 4 degree celsius overnight and then labeled with Donkey anti-Rabbit IgG (H+L) Highly Cross-Adsorbed Secondary... View More

PINK1 Antibody in Immunocytochemistry (ICC/IF)

PINK1 Antibody in Western Blot (WB)

PINK1 Antibody in Western Blot (WB)

PINK1 Antibody in Western Blot (WB)

PINK1 Antibody in Western Blot (WB)

PINK1 Antibody in Western Blot (WB)

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PINK1 Antibody in Immunocytochemistry (ICC/IF) — Immunofluorescence analysis of PINK1 was performed using 70% confluent log phase SH-SY5Y treated with 25 mM Hydrogen Peroxide for 24 hours. The cells were fixed with 4% paraformaldehyde for 10 minutes, permeabilized with 0.1% Triton™ X-100 for 15 minutes, and blocked with 2% BSA for 45 minutes at room temperature. The cells were labeled with PINK1 Polyclonal Antibody (Product # PA5-86941) at 1:200 dilution in 0.1% BSA, incubated at 4 degree celsius overnight and then labeled with Donkey anti-Rabbit IgG (H+L) Highly Cross-Adsorbed Secondary Antibody, Alexa Fluor Plus 488 (Product # A32790, 1:2000 dilution), for 45 minutes at room temperature (Panel a: Green). Nuclei (Panel b:Blue) were stained with ProLong™ Diamond Antifade Mountant with DAPI (Product # P36962). F-actin (Panel c: Red) was stained with Rhodamine Phalloidin (Product # R415, 1:300 dilution). Panel d represents the merged image showing cytoplasmic localization. Panel e represents untreated cells with reduced signal. Panel f represents control cells with no primary antibody to assess background. The images were captured at 60X magnification.

PINK1 Antibody in Western Blot (WB) — Western blot was performed using Anti-PINK1 Polyclonal Antibody (Product # PA5-86941) and a 58 kDa band corresponding to PINK1 was observed across cell lines tested and increased upon Hydrogen Peroxide treatment. Whole cell extracts (20 µg lysate) of HeLa (Lane 1), A549 (Lane 2), SH-SY5Y (Lane 3) and SH-SY5Y treated with 25 mM of Hydrogen Peroxide for 24 hours (Lane 4) were electrophoresed using NuPAGE™ 10% Bis-Tris Protein Gel (Product # NP0301BOX). Resolved proteins were then transferred onto a Nitrocellulose membrane (Product # IB23001) by iBlot® 2 Dry Blotting System (Product # IB21001). The blot was probed with the primary antibody (1:2000 dilution) and detected by chemiluminescence with Goat anti-Rabbit IgG (Heavy Chain) Superclonal™ Recombinant Secondary Antibody, HRP (Product # A27036, 1:8000 dilution) using the iBright FL 1000 (Product # A32752). Chemiluminescent detection was performed using Novex® ECL Chemiluminescent Substrate Reagent Kit (Product # WP20005).

PINK1 Antibody in Western Blot (WB) — Western blot analysis of PINK1 in Lane 1: L02 whole cell lysate (40 µg), Lane 2: A2780 whole cell lysate (40 µg), Lane 3: MCF-7 whole cell lysate (40 µg). Sample was incubated with PINK1 polyclonal antibody (Product # PA5-86941) using a 1:2000 dilution.

PINK1 Antibody in Western Blot (WB) — Knockdown of PINK1 was achieved by transfecting HeLa with PINK1 specific siRNAs (Silencer® select Product # s35166). Western blot analysis (Fig. a) was performed using Whole cell extracts from the PINK1 knockdown cells (lane 3), non-targeting scrambled siRNA transfected cells (lane 2) and untransfected cells (lane 1). The blot was probed with PINK1 Polyclonal Antibody (Product # PA5-86941, 1:2000 dilution) and Goat anti-Rabbit IgG (Heavy Chain) Superclonal™ Recombinant Secondary Antibody, HRP (Product # A27036, 1:8000 dilution). Densitometric analysis of this western blot is shown in histogram (Fig. b). Decrease in signal upon siRNA mediated knock down confirms that antibody is specific to PINK1.

PINK1 Antibody in Western Blot (WB) — Knockout of PINK1 was acheived by CRISPR-Cas9 genome editing in U2OS cells. Western blot analysis of PINK1 was performed by loading 20 µg of U2OS wild type (Lane 1) and U2OS PINK1 KO (Lane 2) whole cell extracts. The blot was probed with Anti-PINK1 Polyclonal Antibody (Product # PA5-86941) using 1:1000 dilution and Goat anti-Rabbit IgG (Heavy Chain), Superclonal™ Recombinant Secondary Antibody, HRP (Product # A27036). Loss of signal of the 45 kDa isoform of PINK1 upon CRISPR mediated knockout (KO) confirms that the antibody is specific to PINK1.

PINK1 Antibody in Western Blot (WB) — Figure 3 FIGURE 3: ZEA blocks cell cycle progression through targeting the potassium channels in the early stages of ZEA treatment. (A) Immunoblotting for KCNJ4, KCNJ12, PINK1 and actin expression after ZEA treatment as well as assessment of cell cycle progression in HCT-8. (B) Immunoblotting for KCNJ4, KCNJ12 and actin expression after ZEA treatment as well as assessment of cell cycle progression in overexpression cell lines. (C) Immunoblotting for KCNJ4, KCNJ12 and actin expression after ZEA treatment as well as their cell cycle in knockout cell lines. (D) ZEA toxin relative concentrations (extracellular and intracellular) in HCT-8 cells exposed to ZEA toxin. (E) Knockout and overexpression of PINK1 and the percentage cellular viability of cell lines after treatment with either 25 or 30 uM of ZEA. (F) Knockout and overexpression of PINK1 and assessment of cell cycle progression. Data are presented as the mean + SEM (n=3). p < 0.05 (*), p < 0.01 (**) and p < 0.001 (***), calculated by Student t-test.

PINK1 Antibody — Antibody specificity was demonstrated by siRNA mediated knockdown of target protein. HeLa cells were transfected with PINK1 siRNA and decrease in signal intensity was observed in Western Blot application using Anti-PINK1 Polyclonal Antibody (Product # PA5-86941). {KD}

PINK1 Antibody — Antibody specificity was demonstrated by CRISPR-Cas9 mediated knockout of target protein. A loss of signal was observed for target protein in PINK1 KO cell line compared to control cell line using Anti-PINK1 Polyclonal Antibody (Product # PA5-86941). {KO}

产品信息

PA5-86941

应用

建议稀释比

已发表文章

免疫印迹 (WB)

1:500-1:2,000

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免疫细胞化学 (ICC/IF)

1:200

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产品规格

种属反应

Human

已发表种属

C. elegans, Human

宿主/亚型

Rabbit / IgG

分类

Polyclonal

类型

Antibody

抗原

Synthetic peptide, corresponding to Human PINK1.

偶联物

Unconjugated Unconjugated Unconjugated

形式

Liquid

浓度

1 mg/mL

纯化类型

Antigen affinity chromatography

保存液

PBS, pH 7.2, with 50% glycerol

内含物

0.02% sodium azide

保存条件

Store at 4°C short term. For long term storage, store at -20°C, avoiding freeze/thaw cycles.

运输条件

Ambient (domestic); Wet ice (international)

产品详细信息

Purity is > 95% by SDS-PAGE.

靶标信息

PINK1 (PTEN-induced putative kinase 1) protects from mitochondrial dysfunction in cells by conferring different autophosphorylation activity to mitochondrial genes. The kinase domain of PINK1 appears to be critical in its protective effects. Mitochondrial proteins are critical for the regulation of cellular energy and adaptation to stress. Particularly in brain cells, mitochondrial demand is extraordinarily high, in order to accommodate aerobic and anaerobic support of high energy processes. When mitochondrial structure and/or function is disrupted in the brain, neurodegenerative diseases like Parkinson's disease are triggered.

仅用于科研。不用于诊断过程。未经明确授权不得转售。

生物信息学

蛋白别名: BRPK; FLJ27236; PINK 1; PINK-1; protein kinase BRPK; PTEN-induced putative kinase protein 1; Serine/threonine-protein kinase PINK1, mitochondrial

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基因别名: BRPK; PARK6; PINK1

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UniProt ID: (Human) Q9BXM7

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Entrez Gene ID: (Human) 65018

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magnesium ion binding protease binding protein kinase activity protein serine/threonine kinase activity protein binding ATP binding calcium-dependent protein kinase activity kinase activity peptidase activator activity ubiquitin protein ligase binding protein kinase B binding C3HC4-type RING finger domain binding non-receptor serine/threonine protein kinase protein modifying enzyme

mitophagy positive regulation of protein phosphorylation regulation of oxidative phosphorylation protein phosphorylation ubiquitin-dependent protein catabolic process response to stress response to oxidative stress mitochondrion organization regulation of hydrogen peroxide metabolic process negative regulation of gene expression regulation of mitochondrion organization positive regulation of peptidase activity macroautophagy positive regulation of macroautophagy negative regulation of macroautophagy phosphorylation protein ubiquitination peptidyl-serine phosphorylation respiratory electron transport chain regulation of protein ubiquitination positive regulation of protein ubiquitination activation of protein kinase B activity positive regulation of synaptic transmission, dopaminergic positive regulation of peptidyl-serine phosphorylation positive regulation of dopamine secretion cellular response to oxidative stress positive regulation of protein dephosphorylation intracellular signal transduction peptidyl-serine autophosphorylation TORC2 signaling positive regulation of I-kappaB kinase/NF-kappaB signaling regulation of protein complex assembly negative regulation of neuron apoptotic process positive regulation of translation negative regulation of JNK cascade protein stabilization positive regulation of sequence-specific DNA binding transcription factor activity positive regulation of ubiquitin-protein transferase activity regulation of mitochondrial membrane potential positive regulation of protein kinase B signaling regulation of proteasomal protein catabolic process cellular response to hypoxia establishment of protein localization to mitochondrion maintenance of protein location in mitochondrion positive regulation of mitochondrial fission positive regulation of release of cytochrome c from mitochondria negative regulation of mitochondrial fission cellular response to toxic substance mitophagy in response to mitochondrial depolarization regulation of synaptic vesicle transport negative regulation of autophagosome assembly positive regulation of mitochondrial electron transport, NADH to ubiquinone regulation of mitophagy negative regulation of mitophagy negative regulation of oxidative stress-induced cell death negative regulation of oxidative stress-induced neuron death regulation of protein targeting to mitochondrion negative regulation of hypoxia-induced intrinsic apoptotic signaling pathway negative regulation of hydrogen peroxide-induced neuron intrinsic apoptotic signaling pathway negative regulation of intrinsic apoptotic signaling pathway in response to hydrogen peroxide positive regulation of cristae formation positive regulation of protein targeting to mitochondrion positive regulation of free ubiquitin chain polymerization regulation of reactive oxygen species metabolic process negative regulation of reactive oxygen species metabolic process positive regulation of ATP biosynthetic process