MA1-058

antibody from Invitrogen Antibodies

Targeting: HTT HD, IT15

Western blot (Supportive data in Antibodypedia)

Immunohistochemistry (Recommended by provider)

Antibody Data

Product number

MA1-058

Provider

Invitrogen Antibodies

Product name

Huntingtin Monoclonal Antibody (4-13)

Antibody type

Monoclonal

Antigen

Synthetic peptide

Description

MA1-058 has been successfully used in Western blot, immunofluorescence, and immunohistochemistry on human and mouse samples. Neoepitope antibodies distinguish smaller cleaved fragments or processed forms of proteins versus the intact full-length or precursor by using a designed peptide purification process to maximize immunoreactivity to a specific cleavage site. Human HTT caspase cleavage sites generate fragment-specific forms of the protein. Caspase-3/7 has been shown to generate cleavage sites at animo acids 513 and 552. Caspase-2 cleaves at amino acid 552 and caspase-6 at amino acid 586. Neo-specific antibody MA1-058 recognizes the 552 cleaved fragment without detecting the full-length form.

Reactivity

Human, Mouse

Host

Mouse

Isotype

IgG

Antibody clone number

13-Apr

Vial size

100 µg

Concentration

1 mg/mL

Storage

-20°C

Western blot

Supportive validation

Submitted by

Invitrogen Antibodies (provider)

Main image

Experimental details

Western blot analysis of over-expressed recombinant HTT fragment lysates was performed by loading 20 µg of lysate per well onto a 4-12% Bis-Tris polyacrylamide gel. Lysate transfected with empty PTET vector was loaded as negative control (Lane 2). Proteins were transferred to a nitrocellulose membrane and blocked with 3% BSA/TBST for at least 1 hour. Membranes were then probed with a neoepitope-specific mouse monoclonal antibody (Product # MA1-058) at a dilution of 1:500 overnight at 4°C on a rocking platform. Membranes were then washed in TBS-0.1%Tween 20 and probed with a goat anti-mouse-HRP secondary antibody at 1:30,000 for at least one hour. Membranes were washed and chemiluminescent detection was performed using SuperSignal West Pico (Product # 34080).

Supportive validation

Submitted by

Invitrogen Antibodies (provider)

Main image

Experimental details

Western blot was performed using Anti-Huntingtin Mouse Monoclonal Antibody (4-13) (Product # MA1-058) and a 62 kDa band corresponding to cleaved fragment of Huntingtin was observed across the tissues tested. Tissue extracts (30 µg lysate) of Mouse Brain (Lane 1), Mouse Kidney (Lane 2) and Mouse Liver (Lane 3) were electrophoresed using NuPAGE™ 3-8% Tris-Acetate Protein Gel (Product # EA0375BOX). 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 (1:1000 dilution) and detected by chemiluminescence with Goat anti-Mouse IgG (H+L) Superclonal™ Recombinant Secondary Antibody, HRP (Product # A28177, 1:4000 dilution) using the iBright FL 1000 (Product # A32752). Chemiluminescent detection was performed using Novex® ECL Chemiluminescent Substrate Reagent Kit (Product # WP20005).