Importin alpha 1 antibody

Overview


	Antigen:	Importin alpha 1
	Antibody Type:	Monoclonal
	Application:	Western Blotting (WB), Immunocytochemistry (ICC), Immunofluorescence (IF)
	Host:	Rat, Mouse, Hamster
	Quantity:	100 µg (1 mg/mL)
	Price:	266,00 € (Plus shipping costs and VAT)
	Order number:	ABIN130690
	Availability:	7 to 10 days until shipment

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Additional Information: Importin alpha 1 antibody

Antigen	Importin alpha 1
Immunogen	This antibody was purified from hybridoma (clone 1A6) supernatant using protein A agarose. This hybridoma was established by fusion of mouse myeloma cell SP2 with WKY/NCrj rat lymphocyte immunized with human recombinant full-length Importin a1
Antibody Type	Monoclonal
Isotype	IgG2a »Matching secondary antibodies
Description	Importing proteins into the nucleus requires at least two sequential steps. First, the target protein is docked onto the nuclear pore complex, followed by an energy-dependent translocation across the nuclear membrane by Ran/TC4, a GTPase. Target proteins contain a short sequence of basic amino acids (Nuclear localization sequence, or NLS). The importin complex is a heterodimer consisting of a ~58 kDa importin a protein and a ~97 kDa importin ß protein. Importin a (also known as karyopherin a, SRP- a , or PTAC58) binds directly to the NLS motif and acts as a NLS receptor. Importin ß (karyopherin ß /PTAC97) is a nuclear transport factor that can also bind to the NLS and enhance target protein binding. The importin ß-target complex is translocated through the nuclear pore via interactions with a series of nucleoporins. Importin ß also binds and inhibits Ran, consequently providing a mechanism of nuclear transport termination. Several importin a isotypes have been identified, each exhibiting differential recognition and nuclear transport, probably via preferential binding to a particular NLS.
Clone	1A6
Host	Rat, Mouse, Hamster
Specificity	This antibody reacts with Importin a1 (~60 kDa) on Western blotting.

Application Details: Importin alpha 1 antibody

Application	Western Blotting (WB), Immunocytochemistry (ICC), Immunofluorescence (IF)
Application Notes	SDS-PAGE & Western Blotting 1) Wash the cells 3 times with PBS and suspend with 10 volumes of cold Lysis buffer (50 mM Tris-HCl, pH 7.2, 250 mM NaCl, 0.1% NP-40, 2 mM EDTA, 10% glycerol) containing appropriate protease inhibitors. Incubate it at 4 o C with rotating for 30 minutes, then sonicate briefly (up to 10 seconds). 2) Centrifuge the tube at 12,000 x g for 10 minutes at 4 o C and transfer the supernatant to another tube. Measure the protein concentration of the supernatant and add the cold Lysis buffer to make 8 mg/mL solution. 3) Mix the sample with equal volume of Laemmli’s sample buffer. 4) Boil the samples for 2 minutes and centrifuge. Load 10 µL of the sample per lane in a 1 mm thick SDS-polyacrylamide gel for electrophoresis. 5) Blot the protein to a polyvinylidene difluoride (PVDF) membrane at 1 mA/cm 2 for 1 hour in a semi-dry transfer system (Transfer Buffer - 25 mM Tris, 190 mM glycine, 20% MeOH). See the manufacture's manual for specific transfer procedure. 6) To reduce nonspecific binding, soak the membrane in 10% skimmed milk (in PBS, pH 7.2) for 1 hour at room temperature, or overnight at 4 o C. 7) Incubate the membrane with primary antibody diluted with PBS, pH 7.2 containing 1% skimmed milk as suggested in the Applications for 1 hour at room temperature. (The optimal antibody concentration will depend on the experimental conditions.) 8) Wash the membrane with PBS (5 minutes x 6 times). 9) Incubate the membrane with the 1:5,000 HRP conjugated anti-Rat IgG (H+L) (Immunotech, code IM-0825) diluted with 1% skimmed milk (in PBS, pH 7.2) for 1 hour at room temperature. 10) Wash the membrane with PBS (5 minutes x 6 times). 11) Wipe excess buffer from the membrane, then incubate it with appropriate chemiluminescence reagents for 1 minute. Remove extra reagent from the membrane by dabbing with a paper towel, and seal it in plastic wrap. 12) Expose to X-ray film in a dark room for 5 minutes. Develop the film as usual. The conditions for exposure and development may vary. Positive controls for Western blotting, HeLa, Jurkat, Raji, HL60, A431, NIH/3T3, CHO, PC12 Immunofluorescence microscopy 1) Culture the cells in the appropriate condition on a glass slide (for example, spread 10 4 of HeLa cells for one slide, then incubate in a CO 2 incubator for one night.) 2) Fix the cells by immersing the slide in PBS containing 4 % Paraformaldehyde for 10 minutes on ice. 3) Immerse the slide in PBS containing 0.1% TritonX-100 for 10 minutes at room temperature. 4) Add the primary antibody diluted with PBS as suggested in the Applications onto the cells and incubate for 1 hour at room temperature (Optimization of antibody concentration and incubation conditions is recommended.) 5) Prepare a wash container such as a 500 mL beaker with a stirrer. Then wash the cultured cells on the glass slide by soaking the slide with a plenty of PBS in the wash container for 5 minutes. Be careful not to touch the cells. Repeat washes once more. 6) Add the secondary antibody (1:100 FITC conjugated anti-mouse IgG/code 238) onto the cells. Incubate for 1 hour at room temperature. Keep out light using aluminum foil. 7) Wash the slide in a plenty of PBS as in the step 5). 8) Wipe excess liquid from slide but be careful not to touch the cells. Never permit the cells to dry. 9) Promptly add mounting medium onto the slide, then put a cover slip on it. Positive control for Immunocytochemistry, HeLa 050225-1 Im m unocytochem ical detection of Im portin a1 on 4% P FA fixed HeLacells w ith D 168-3. Im m unocytochem ical detection of Im portin a1 on 4% P FA fixed HeLacells w ith D 168-3. Western blotting: 1µg / mL for chemiluminescence detection system. Immunoprecipitation: Not tested. Immunohistochemistry: Not recommended. Immunocytochemistry: 5-10 µg / mL Protocol. This antibody has also been successfully used in immunofluorescence staining (Kamikubo, Y. et al., 2004). Species Cross
Concentration	1 mg/mL
Buffer	100 µg IgG in 100 µL P BS containing 50% glycerol, pH 7.2. Contains no preservatives.
Restrictions	For Research Use Only

Publications: Importin alpha 1 antibody

Publications

Sakaguchi, Miyamoto, Yoneda et al.:
"Generation of a rat monoclonal antibody specific for importin alpha3/Qip1."
In: Hybridoma and hybridomics , Vol. 22, Issue 6, pp. 397-400, 2003/01 (PubMed)

Kamikubo, Sakaguchi, Shikata et al.:
"Specific monoclonal antibody against nuclear import factor, importin alpha1/Rch1."
In: Hybridoma and hybridomics , Vol. 23, Issue 5, pp. 301-4, 2005/01 (PubMed)

Images: Importin alpha 1 antibody

Western blot analysis of Importin ? 1 expression in Jurkat cells (1), HeL a cells (2), Raji cells (3), HL60 cells (4), A431 cells (5), L 5178Y cells (6), WR19L cells (7), NIH-3T3 cells (8), CHO cells (9), and PC12 cells (10) using D 168-3.

Image for Importin alpha 1 antibody (1)

Immunocytochemical detection of Importin ?1 on 4% PFA fixed H eLa cells with ABIN130690.

Image for Importin alpha 1 antibody (2)

External Information: Importin alpha 1 antibody

Google Scholar	Find more references for clone 1A6 on Google Scholar™
EMBL Harvester	Search human, mouse or rat genome for antigen Importin alpha 1 (Bioinformatic Harvester (EMBL))