Alkaline Phosphatase 抗体 (YA629)

Alkaline Phosphatase Antibody (YA629) is a Rabbit-derived and non-conjugated IgG monoclonal antibody, targeting to Alkaline Phosphatase.

Alkaline Phosphatase（ALP）是一种碱性磷酸酶，能够代谢多种磷酸化合物，在骨骼矿化和适应性产热中起关键作用。该酶具有广泛的底物特异性，可水解多种化合物，但天然底物主要包括二磷酸盐（无机焦磷酸盐，PPi）、5’-磷酸吡哆醛（PLP）和N-磷酸肌酸。通过水解细胞外二磷酸盐（一种强效矿化抑制剂）生成磷酸盐，ALP促进羟基磷灰石晶体形成并提高无机磷酸盐浓度，从而在骨骼和牙齿矿化中发挥重要作用（By similarity）。与PHOSPHO1在骨骼矿化中具有非冗余功能：PHOSPHO1介导基质小泡（MVs）中羟基磷灰石结晶的起始，而ALPL/TNAP则催化细胞外基质中羟基磷灰石结晶的扩展。此外，ALP还能催化PLP去磷酸化为吡哆醛（PL），这是维生素B6的可运输形式，为大脑中多种神经递质合成酶提供必需辅因子。该酶还能逐步介导ATP降解为腺苷，从而调节嘌呤受体配体的可用性（By similarity），并能够去磷酸化微生物产物（如脂多糖LPS）和其他磷酸化小分子（如聚肌胞苷酸poly I:C）。在适应性产热中，ALP作为无效肌酸循环的关键调节因子，通过介导N-磷酸肌酸水解启动肌酸去磷酸化和磷酸化的无效循环，将N-磷酸肌酸的高能电荷以热量形式耗散。

Free

P05186

Predicted band size: 57 kDa；Observed band size: 75 kDa

Protein A affinity purified.

Cell membrane, Mitochondrion membrane, Mitochondrion intermembrane space, Extracellular vesicle membrane.

Non-conjugated

Unmodified

AB_3102261

Tags & Cell Markers

Primary Antibody; Recombinant Rabbit Monoclonal Antibody

Recombinant, Monoclonal

Rabbit

Human, Mouse

WB: 1:5000; IHC-P: 1:11000-1:8000; IF-Tissue: 1:200

• 
  Western blot analysis of extracts from A549 (lane 2(20μg) , Hela(lane 3(20μg) and Saos-2(lane 4(20μg) using Alkaline Phosphatase(HY-P80012) Rabbit mAb. Proteins were transferred to a PVDF membrane and blocked with 5% non-fat milk in TBST for 2 hour at room temperature. The primary antibody (1/1000) and Loading control antibody (Beta Actin, HY-P80438, 1/10000) was used in 5% non-fat milk in TBST at 4°C overnight. Goat Anti-Mouse IgG-HRP Secondary Antibody (1/10000) was used for 1 hour at room temperature.
• 
  Immunocytochemistry analysis of Hela cells labeling Alkaline Phosphatase with Alkaline Phosphatase Antibody (HY-P80012)at 1/50 dilution. Cells were fixed in 4% paraformaldehyde for 15 minutes at room temperature, permeabilized with 0.1% Triton X-100 for 10 minutes at room temperature, then blocked with QuickBlock™ Blocking Buffer for Immunol Staining for 10 min at room temperature. Cells were then incubated with Alkaline Phosphatase Antibody (HY-P80012) at 1/50 dilution in QuickBlock™ Blocking Buffer for Immunol Staining at 4 ℃. Alexa Fluor® 488-conjugated AffiniPure Goat Anti-Rabbit IgG H&L（HY-P8002, Green) was used as the secondary antibody at 1/1,000 dilution. PBS instead of the primary antibody was used as the secondary antibody only control. The Nuclear counterstain was DAPI (Blue).
• 
  Immunocytochemistry analysis of A549 cells labeling Alkaline Phosphatase with Alkaline Phosphatase Antibody (HY-P80012) at 1/100 dilution. Cells were fixed in 4% paraformaldehyde for 15 minutes at room temperature, permeabilized with 0.1% Triton X-100 for 10 minutes at room temperature, then blocked with QuickBlock™ Blocking Buffer for Immunol Staining for 10 min at room temperature. Cells were then incubated with Alkaline Phosphatase Antibody (HY-P80012) at 1/100 dilution in QuickBlock™ Blocking Buffer for Immunol Staining at 4 ℃. Alexa Fluor® 488-conjugated AffiniPure Goat Anti-Rabbit IgG H&L（HY-P8002, Green) was used as the secondary antibody at 1/1,000 dilution. PBS instead of the primary antibody was used as the secondary antibody only control. The Nuclear counterstain was DAPI (Blue).
• 
  Immunohistochemical analysis of paraffin-embedded Mouse liver tissue using Alkaline Phosphatase Antibody. The section was pre-treated using heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0) for 8 minutes. The tissues were blocked in QuickBlock for 20 minutes at room temperature, washed with ddH2O and PBS, and then probed with the primary antibody (HY-P80012, 1/2000) in 4℃ overnight. The detection was performed using an HRP conjugated compact polymer system. DAB was used as the chromogen. Tissues were counterstained with hematoxylin and mounted with DPX.
• 
  Immunohistochemical analysis of paraffin-embedded Mouse liver tissue using Alkaline Phosphatase Antibody. The section was pre-treated using heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0) for 8 minutes. The tissues were blocked in QuickBlock for 20 minutes at room temperature, washed with ddH2O and PBS, and then probed with the primary antibody (HY-P80012, 1/2000) in 4℃ overnight. The detection was performed using an HRP conjugated compact polymer system. DAB was used as the chromogen. Tissues were counterstained with hematoxylin and mounted with DPX.
• 
  Immunohistochemical analysis of paraffin-embedded human Gastric Carcinoma‌ tissue using Alkaline Phosphatase antibody was performed. The section was pretreated using high-temperature and high-pressure mediated EDTA antigen retrieval buffer (pH 9.0), for 5 minutes. The tissues were incubated with primary antibody (HY-P80012, 1:1000 dilution) at room temperature for 60 minutes. Detection was performed using an HRP conjugated compact polymer system. DAB was used as the chromogen. The tissues were counterstained with hematoxylin and mounted with neutral balsam mounting medium.
• 
  Immunohistochemical analysis of paraffin-embedded human Liver Cancer tissue using Alkaline Phosphatase antibody was performed. The section was pretreated using high-temperature and high-pressure mediated EDTA antigen retrieval buffer (pH 9.0), for 5 minutes. The tissues were incubated with primary antibody (HY-P80012, 1:1000 dilution) at room temperature for 60 minutes. Detection was performed using an HRP conjugated compact polymer system. DAB was used as the chromogen. The tissues were counterstained with hematoxylin and mounted with neutral balsam mounting medium.
• 
  Tyramide signaling amplification based immunofluorescence was performed on paraffin-embedded human ovarian carcinoma (sample 1) tissue using Alkaline Phosphatase antibody. Antigen retrieval was performed in EDTA buffer pH 9.0 (95 °C, 20 min) followed by cooling to RT. Then incubated with primary antibody (HY-P80012, 1:1000 dilution) at room temperature for 60 minutes and HRP conjugated secondary antibody for 10 minutes. Fluorescence was then developed with TSA520 . The tissues were counterstained with DAPI and mounted with Anti-fade mounting medium.
• 
  Tyramide signaling amplification based immunofluorescence was performed on paraffin-embedded human ovarian carcinoma (sample 2) tissue using Alkaline Phosphatase antibody. Antigen retrieval was performed in EDTA buffer pH 9.0 (95 °C, 20 min) followed by cooling to RT. Then incubated with primary antibody (HY-P80012, 1:1000 dilution) at room temperature for 60 minutes and HRP conjugated secondary antibody for 10 minutes. Fluorescence was then developed with TSA520 . The tissues were counterstained with DAPI and mounted with Anti-fade mounting medium.

WB, IHC-P, IF-Tissue

液体

Supplied in 1*TBS (pH7.4), 0.05% BSA and 40% Glycerol. Preservative: 0.05% Sodium Azide.

Stored at -20°C for 1 year. Avoid repeated freeze / thaw cycles.

Shipping with blue ice.

IgG

Endogenous

Synthetic peptide corresponding to Human Alkaline Phosphatase.AA range:18-50.