Analysis Note

ControlNormal human cerebral cortex lysate, mouse brain cortex samples from HD or wild type miceHEK293 lysates.

Application

Research Sub CategoryNeurodegenerative Diseases

Research CategoryNeuroscience

Immunohistochemistry:
1:50-1:100 of a previous lot using ABC on 4% paraformaldehyde fixed tissue. Suggested dilution buffer is PBS containing 3% BSA. The antibody works on paraffin embedded tissue sections.
Suggested dilution buffer is PBS containing 3% BSA. The antibody works on paraffin embedded tissue sections. Yu, Z et al (2002) Hum. Mole. Genetics 11(8):905-914. (http://hmg.oxfordjournals.org/cgi/content/full/11/8/905) for good IHC methods and photos of mEM48 on rodent tissues with human transgenic material.

Immunocytochemistry:
light 4% PFA fixation followed by 0.1% triton X-100 incubation prior to blocking is suggested.
A previous lot of this antibody was used in IC.

Western blot:
1:50-1:500 using ECL depending on the level of mutant protein. Suggested dilution buffer is PBS containing 3% BSA or PBS containing 5% non-fat milk.
Nuclear fraction preparations enhance signals; monomeric protein ~80kDa; aggregates are common which can be >200kDa in size.

Optimal working dilutions must be determined by the end user.

Detect Huntingtin Protein using this Anti-Huntingtin Protein Antibody, clone mEM48 validated for use in IC, IH & WB.

Disclaimer

Unless otherwise stated in our catalog or other company documentation accompanying the product(s), our products are intended for research use only and are not to be used for any other purpose, which includes but is not limited to, unauthorized commercial uses, in vitro diagnostic uses, ex vivo or in vivo therapeutic uses or any type of consumption or application to humans or animals.

General description

Huntington disease (HD) is a hereditary, progressive, neurodegenerative ailment characterized by personality changes, motor impairment and subcortical dementia. The molecular basis of the disease involves the expansion of the trinucleotide CAG, coding for polyglutamine in the first exon of a chromosome four gene (4p16.3), which normally produces a widely expressed 3136 a.a. (~350 kDa) protein huntingtin with unclear function. The protein is found in the perinuclear region along with microtubules, and in the centrosomal region along with gamma-tubulin. Huntingtin is necessary for neuronal survival and is involved in synaptic vesicle trafficking, microtubule binding and may also have a role in apoptosis. In the HD condition, neuronal cells with the mutant form of huntingtin possess intranuclear aggregations of the N-terminal fragment, causing damaging inclusions in perinuclear locations and striatal neuron cell death. Wild-type huntington and anti-huntingtin reduce aggregation and cellular toxicity of the mutant huntingtin form in mammalian cell models of HD. Huntingtin is known to interact with GAPDH, HAP-1, SP1 and TAFII130.

Immunogen

GST fusion protein from the first 256 amino acids from human huntingtin with the deletion of the polyglutamine tract.

Legal Information

CHEMICON is a registered trademark of Merck KGaA, Darmstadt, Germany

Other Notes

Concentration: Please refer to the Certificate of Analysis for the lot-specific concentration.

Physical form

Culture Supernatant mouse monoclonal IgG containing no preservative.

Unpurified

Quality

Routinely evaluated by Western Blot on HEK293 lysates.

Western Blot Analysis:
1:1000 dilution of this lot detected Huntingtin Protein on 10 µg of HEK293 lysates.

Specificity

Reacts with human huntingtin protein (both native and recombinant protein). MAB5374 reacts with mutant huntingtin in patients and in transgenic animals that express different numbers of repeats (from 82 to 150 glutamines). Thus, it should recognize different forms of mutant huntingtin.

Storage and Stability

Stable for 6 months at -20°C in undiluted aliquots from date of receipt.
Handling Recommendations: Upon receipt, and prior to removing the cap, centrifuge the vial and gently mix the solution. Aliquot into microcentrifuge tubes and store at -20°C. Avoid repeated freeze/thaw cycles, which may damage IgG and affect product performance.

Target description

over 200 kDa