Actin antibody

Catalog No. ABIN3181106

Product Details anti-Actin Antibody

Target: Actin

Reactivity: Plant

Host: Mouse

Clonality: Monoclonal

Conjugate: This Actin antibody is un-conjugated

Application: Western Blotting (WB)

Specificity: The antibody detects endogenous plant actin protein.

Characteristics: Mouse Monoclonal to Plant actin.

Purification: The antibody was affinity-purified from mouse ascites by affinity-chromatography using specific immunogen.

Clone: Q30

Isotype: IgG1

Application Details

Application Notes: WB 1:2000-5000

Restrictions: For Research Use only

Independent Validation (WB)

Validation #102343 (Western Blotting)

by: Department of Horticultural genetics and Biotechnology, Mediterranean Agronomic Institute of Chania

No.: #102343

Date: 12/19/2017

Antigen: Actin

Lot Number: B3401

Method validated: Western Blotting

Positive Control: protein extraction from olive root tissue and tomato flower tissue

Notes:

Passed. ABIN3181106 specifically recognizes the actin in both tomato flower and olive roots samples, providing a clear band at the detection with the secondary one.

Validation Images

Detection of Actin in tomato flower from wt plants and different RNAi lines (A) and olive root protein extracts from different cultivars (B) using ABIN3181106 diluted 1:2500 and 1:1000 respectively.

Full Methods

Primary Antibody: ABIN3181106

Secondary Antibody: HRP conjugated rabbit anti-mouse IgG (Thermo Scientific, PA1-28902, OK1782904)

Full Protocol:

• Lyse protein extraction from 500mg olive root tissue and 80mg from tomato flower tissue in 240µl cold lysis buffer (1M Tris-Cl pH6.8, 10% SDS, 14.3M β-Merc, 0.5N EDTA, PMSF 0.5mM and 100µl proteinase inhibitor (Sigma-Aldrich, P9599-1ML)).
• Determine total protein content of the lysates using protein assay (Bradford curve).
• Denature 0.7μg/μl of total protein for 5min at 95°C 1:4 (buffer:protein) in Laemmli SDS sample buffer (Tris-Cl pH6.8, 2% SDS, 10% glycerol, 1%β-mercaptoethanol, 12.5mM EDTA, 0.02% bromophenol blue) and subsequently separate them on a freshly cast denaturing SDS-PAGE gel (30% Bis-Acrylamide, 1.5M Tris pH8.8, 10% SDS, 10% APS and TEMED) in an Bio-Rad Mini-PROTEAN 3 cell systemfor 2h at 100V, 200mA.
• Transfer proteins onto an Immobilon-P membrane (Millipore, IPVPH0010, K5CA2260G) with a Western blotting system for 1h at 100V.
• Block the membrane with 5% non-fat milk in 1x TBST for 1h at RT.
• Incubate membrane with anti-Actin antibody (antibodies-online, ABIN3181106, lot B3401) diluted 1:2500 (Flower samples) or 1:1000 (root samples) in 5% non-fat milk in 1x TBST ON at 4°C.
• Wash membrane 3x for 5min with 1x TBST.
• Incubate membrane with secondary HRP conjugated rabbit anti-mouse IgG (Thermo Scientific, PA1-28902, OK1782904) diluted 1: 5000 (flower samples) and 1:2000 (root samples) in 5% non-fat milk in 1x TBSTfor 1h at RT.
• Wash membrane 3x for 5min with 1X TBST.
• Reveal protein bands using super signal west pico (Thermo-Scientific, 34080, QF218483) on a Super RX Fuji Medical X-RAY (18cm x 24cm) film.

Experimental Notes:

The actin antibody ABIN3181106 reveals the actin protein of the expected molecular weight of antigen in lysates of tomato flower and olive root samples.

Handling

Format: Liquid

Concentration: 1 mg/mL

Buffer: Liquid in PBS containing 50 % glycerol, 0.5 % BSA and 0.02 % sodium azide.

Preservative: Sodium azide

Precaution of Use: This product contains Sodium azide: a POISONOUS AND HAZARDOUS SUBSTANCE which should be handled by trained staff only.

Handling Advice: Avoid repeated freeze/thaw cycles.

Storage: -20 °C

Storage Comment: Store at -20°C.

References for anti-Actin antibody (ABIN3181106)

Hoffmann, Dietrich, Herrmann, Schacht, Albrecht, Methner: "Dimethyl Fumarate Induces Glutathione Recycling by Upregulation of Glutathione Reductase." in: Oxidative medicine and cellular longevity, Vol. 2017, pp. 6093903, (2017) (PubMed).

Target Details for Actin

Target: Actin

Abstract: Actin Products