anti-p62/ SQSTM1 (C-terminus) + positive western blot control
|Quantity||100 µl anti-p62 + 50 µg western blot control|
|Purification||Cell culture lysate, Stabilized antiserum|
|Reactivity||Bovine, Human, Mouse, Rat|
|Storage||Antibody: short term at 2-8°C; long term in aliquots at -20°C; western blot control: lyophilized at 2-8°C, reconstituted at -20°C; avoid freeze/thaw cycles for both components|
|Intended use||Research use only|
|Immunogen||C-terminal domain (20 amino acids: C-NYD IGA ALD TIQ YSK HPP PL) of human p62 protein, coupled to KLH. This peptide sequence is identical in human, monkey, bovine, mouse, and rat.|
|Formulation||antibody: contains 0.09% sodium azide; western blot control: lyophilized, reconstitute in 50 µl 1 x SDS buffer (lysis buffer composition: PBS + Pefablock)|
|Note||Centrifuge prior to opening|
|Tested applications||Tested dilutions|
|Immunohistochemistry (IHC) - frozen||anti-p62: 1:100-1:600|
|Immunohistochemistry (IHC) - paraffin||anti-p62: 1:100-1:600 (microwave treatment recommended)|
|Western Blot (WB)||anti-p62: 1:1,000-1:3,000; western blot control: 10 µg total protein per lane|
The anti-p62 antibody is useful for research in ubiquitin-associated degradation and autophagy and for detection of neurofibrillary tangles in the brain of Alzheimer disease patients, in Parkinson diseases and various chronic liver diseases.
Human 62 kDa (p62) protein, is present in intracytoplasmic inclusions (e.g. hyaline bodies) of hepatocellular carcinoma.
p62 protein (also described as ubiqutin-binding protein; sequestosome 1; SQSTM1) has been found in many tissues and cells, including lymphoid cells, serving probably a common cellular signal transduction mechanism (e.g. ubiquitin-associated degradation and autophagy).
The antiserum stains also neurofibrillary tangles in the brain of patients suffering from Alzheimer's disease.
The GP62-C antibody is knockout validated (Waguri & Komatsu, 2009).
Positive western blot control:
Whole Cell Lysate from PLC/PRF/5 human Hepatoma cell line.
PLC whole cell lysate was prepared by homogenization in PBS containing Pefablock.
Protein concentration was determined using Bradford assay.
Q & A's
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- PVDF membranes show better results than nitrocellulose (higher capacity, allows for more stringent washing conditions in case of background problems).
- Use freshly prepared blocking solution (e.g. 5% nonfat dry milk, 0.05% Tween 20), block for at least 1 h at room temperature.
- Use the antibody in a higher dilution, but prolong incubation time and exposure time.
- Always use a fresh aliquot of the antibody.
- Do not repeatedly freeze the antibody (eventually centrifuge shortly after thawing to remove cryo-precipitates).
- Include an additional washing step.
You might also try more stringent wash conditions, e.g. add 0.5 M NaCl to the wash buffer.
- Always use a fresh aliquot of secondary antibody.
- In case you use ECL most the guinea pig antibody should be diluted further in order to get rid of the background.
- Supernatant and supernatant concentrate: This format contains hybridoma cell culture supernatant. The antibody is not purified and the antibody concentration is not determined. The antibody concentration may vary from lot to lot. Therefore we recommend to titrate the optimal concentration for the application used for each new lot.
- Lyophilized, purified: This format contains purified antibody in lyophilized form. The reconstitution of this antibody is described in the datasheet. The buffer composition after reconstitution is also mentioned on the datasheet.
- Liquid, purified: This format contains purified antibody in liquid format. The concentration is mentioned on the datasheet.
- Prediluted, purified: This format contains purified antibody in liquid format. Most antibodies in this format are diluted to be ready-to-use for IHC with standard tissue. But some antibodies of this format need further dilution for IHC. This is mentioned on the datasheet.
- Whole cell lysate from PLC human hepatoma cell line (PLC/PRF/5)
- Validated and tested western blot control for: anti-p62 / SQSTM1 (C-terminus) guinea pig polyclonal antibody, Cat. No. GP62-C