anti-Phakinin guinea pig polyclonal, serum
- Guinea pig polyclonal
- Suitable for IHC and WB
- Reacts with bovine and human
|Storage||Short term at 2-8°C; long term storage in aliquots at -20°C; avoid freeze/thaw cycles|
|Intended use||Research use only|
|Immunogen||Phakinin purified from calf lens|
|Formulation||Contains 0.09% sodium azide and 0.5% BSA|
|UniprotID||Q28177 (Bovine),Q13515 (Human)|
|Synonym||Phakinin, 49 kDa cytoskeletal protein, Beaded filament structural protein 2, Lens fiber cell beaded filament protein CP 47, CP47, Lens fiber cell beaded filament protein CP 49, CP49, Lens intermediate filament-like light, LIFL-L, BFSP2|
|Note||Centrifuge prior to opening|
|Tested applications||Tested dilutions|
|Immunohistochemistry (IHC) - frozen||1:100|
|Western Blot (WB)||1:5,000|
Specific detection of phakinin (also described as CP49; Mr 47,000 polypeptide), intermediate filament protein detected in lens.
Specific note: the antiserum cross-reacts with filensin (also described as CP115; Mr 115,000 polypeptide also present in lens).
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.
The supernatant format contains FCS proteins from cell culture medium supplemented with FCS.
The serum antibodies contain other proteins present in serum.
Most of our liquid antibodies and reconstituted lyophilized antibodies may be stored for short term storage (up to 3 month) at 2-8°C. For long term storage we recommend to store the antibody at -20°C in aliquots. Please avoid freeze and thaw cycles.
Most of our conjugated antibodies should be stored at 2-8°C.
The individual storage conditions are mentioned on the datasheet.
- Methanol/ acetone fixation: Immerse slide in precooled (-20°C) methanol for 5 min, immerse in precooled (-20°C) acetone for 30-60 sec, let specimen air dry before antibody incubation.
- Methanol/ acetone fixation plus detergent permeabilization: After methanol/ acetone fixation and air-drying dip slide either in a solution containing 0.1-0.2% Triton X-100 in PBS or in 0.1% saponin in PBS for 1-5 min at room temperature (enhances accessibility of many cytoskeletal antigens).
- Air-drying of the section.
- Block with the serum of the species in which the secondary antibody was raised for 30 min.
- Incubation with 1st antibody 1 h at RT in moist chamber.
- Wash 3x with PBS.
- Incubation with appropriate fluorescent secondary antibody, 30-60 min at RT.
- Wash 3x with PBS.
- Immerse shortly into ethanol.
- Let air dry.
- Cover with mounting medium.
In guinea pigs the antibody concentration in serum varies from 10 to 20 mg/ml; specific antibodies represent normally about 0.1-1% of total IgG. Total protein concentration varies from 40 to 65 mg/ml, with the main constituent (about 60%) being albumin.