559304
Anti-SAPK/JNK Rabbit pAb
liquid, Calbiochem®
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About This Item
Code UNSPSC :
12352203
Nomenclature NACRES :
NA.41
Produits recommandés
Source biologique
rabbit
Niveau de qualité
Forme d'anticorps
affinity isolated antibody
Type de produit anticorps
primary antibodies
Clone
polyclonal
Forme
liquid
Ne contient pas
preservative
Espèces réactives
mouse, rat, human
Fabricant/nom de marque
Calbiochem®
Conditions de stockage
OK to freeze
avoid repeated freeze/thaw cycles
Isotype
IgG
Conditions d'expédition
wet ice
Température de stockage
−20°C
Modification post-traductionnelle de la cible
unmodified
Informations sur le gène
human ... MAPK8(5599)
Description générale
Protein A and immunoaffinity purified rabbit polyclonal antibody. Recognizes the ~54 kDa SAPK/JNK protein.
Recognizes the ~54 kDa SAPK/JNK protein in uv-treated HEK293 cells.
This Anti-SAPK/JNK Rabbit pAb is validated for use in Immunoblotting, Immunocytochemistry for the detection of SAPK/JNK.
Immunogène
Human
a full-length, recombinant, human p54 SAPK/JNK2 fusion protein
Application
Immunoblotting (1:1000)
Immunocytochemistry (1:200)
Immunocytochemistry (1:200)
Avertissement
Toxicity: Standard Handling (A)
Forme physique
In 150 mM NaCl, 10 mM HEPES, 50% glycerol, 0.01% BSA, pH 7.5.
Reconstitution
Following initial thaw, aliquot and freeze (-20°C).
Remarque sur l'analyse
Positive Control
UV treated HEK293 cells
UV treated HEK293 cells
Autres remarques
Gupta, S., et al. 1996. EMBO J.15(11), 2760.
Coso, O.A., et al. 1995. Cell81, 1137.
Derijard, B., et al. 1994. Cell76, 1025.
Kyriakis, J.M., et al. 1994. Nature369, 156.
Hibi, M., et al. 1993. Genes Dev.7, 2135.
Kyriakis, J.M. and Avruch, J. 1990. J. Biol. Chem.265, 17355.
Coso, O.A., et al. 1995. Cell81, 1137.
Derijard, B., et al. 1994. Cell76, 1025.
Kyriakis, J.M., et al. 1994. Nature369, 156.
Hibi, M., et al. 1993. Genes Dev.7, 2135.
Kyriakis, J.M. and Avruch, J. 1990. J. Biol. Chem.265, 17355.
Recognizes SAPK/JNK regardless of the phosphorylation state. Variables associated with assay conditions will dictate the proper working dilution.
Recommended Protocol for Immunoblotting
Solutions and Reagents
•Transfer Buffer: 25 mM Tris base, 0.2 M glycine, 20% methanol, pH 8.5.
•SDS Sample Buffer: 62.5 mM Tris-HCl, pH 6.8, 2% SDS, 10% glycerol, 50 mM DTT, 0.1% bromphenol blue.
•10X TBS (Tris-buffered saline): To prepare 1 liter, 24.2 g Tris base, 80 g NaCl, adjust pH to 7.6 with HCl. Dilute 1:10 for use.
•Blocking Buffer: 1X TBS, 0.1% Tween®-20 detergent with 5% non-fat dry milk.
•Primary Antibody Dilution Buffer: 1X TBS, 0.1% Tween-20 detergent with 5% BSA
•Wash Buffer (TBST): 1X TBS, 0.1% Tween-20 detergent
Blotting Membrane
Nitrocellulose or PVDF membranes may be used.
Protein Blotting
A general protocol for sample preparation using 2x106 293 cells per well in a 6-well plate is as follows:
1. Aspirate media. Treat cells by adding fresh media containing regulator for desired time.
2. Aspirate media from cultures; wash cells with PBS; aspirate.
3. Lyse cells by adding 100 µl of SDS Sample Buffer and immediately scrape the cells off the plate and transfer the extract to a microfuge tube. Keep on ice.
4. Sonicate for 2 s to shear DNA and reduce sample viscosity.
5. Heat sample to 95-100°C for 5 min. Cool on ice.
6. Microcentrifuge for 5 min.
7. Load 20 µl onto SDS-PAGE gel (10 cm x 10 cm).
8. Electrotransfer to nitrocellulose membrane.
As controls, we recommend using 20 µl lysate from UV treated HEK293 cell.
Membrane Blocking, Gel and Antibody Incubations
1. After transfer, wash membrane with 25 ml TBS for 5 min at room temperature.
2. Incubate membrane in 25 ml Blocking Buffer for 1-3 h at room temperature or overnight at 4°C.
3. Wash 3 times for 5 min each with 15 ml TBST.
4. Incubate membrane and primary antibody (at the appropriate dilution) in 10 ml Primary Antibody Dilution Buffer with gentle agitation overnight at 4°C.
5. Wash 3 times for 5 min each with 15 ml TBST.
6. Incubate membrane with conjugated secondary antibody at the appropriate dilution in 10 ml Blocking Buffer with gentle agitation for 1 h at room temperature.
7. Wash membrane as in step 5.
Detection of Proteins
Chemiluminescence.
Recommended Protocol for Immunoblotting
Solutions and Reagents
•Transfer Buffer: 25 mM Tris base, 0.2 M glycine, 20% methanol, pH 8.5.
•SDS Sample Buffer: 62.5 mM Tris-HCl, pH 6.8, 2% SDS, 10% glycerol, 50 mM DTT, 0.1% bromphenol blue.
•10X TBS (Tris-buffered saline): To prepare 1 liter, 24.2 g Tris base, 80 g NaCl, adjust pH to 7.6 with HCl. Dilute 1:10 for use.
•Blocking Buffer: 1X TBS, 0.1% Tween®-20 detergent with 5% non-fat dry milk.
•Primary Antibody Dilution Buffer: 1X TBS, 0.1% Tween-20 detergent with 5% BSA
•Wash Buffer (TBST): 1X TBS, 0.1% Tween-20 detergent
Blotting Membrane
Nitrocellulose or PVDF membranes may be used.
Protein Blotting
A general protocol for sample preparation using 2x106 293 cells per well in a 6-well plate is as follows:
1. Aspirate media. Treat cells by adding fresh media containing regulator for desired time.
2. Aspirate media from cultures; wash cells with PBS; aspirate.
3. Lyse cells by adding 100 µl of SDS Sample Buffer and immediately scrape the cells off the plate and transfer the extract to a microfuge tube. Keep on ice.
4. Sonicate for 2 s to shear DNA and reduce sample viscosity.
5. Heat sample to 95-100°C for 5 min. Cool on ice.
6. Microcentrifuge for 5 min.
7. Load 20 µl onto SDS-PAGE gel (10 cm x 10 cm).
8. Electrotransfer to nitrocellulose membrane.
As controls, we recommend using 20 µl lysate from UV treated HEK293 cell.
Membrane Blocking, Gel and Antibody Incubations
1. After transfer, wash membrane with 25 ml TBS for 5 min at room temperature.
2. Incubate membrane in 25 ml Blocking Buffer for 1-3 h at room temperature or overnight at 4°C.
3. Wash 3 times for 5 min each with 15 ml TBST.
4. Incubate membrane and primary antibody (at the appropriate dilution) in 10 ml Primary Antibody Dilution Buffer with gentle agitation overnight at 4°C.
5. Wash 3 times for 5 min each with 15 ml TBST.
6. Incubate membrane with conjugated secondary antibody at the appropriate dilution in 10 ml Blocking Buffer with gentle agitation for 1 h at room temperature.
7. Wash membrane as in step 5.
Detection of Proteins
Chemiluminescence.
Informations légales
CALBIOCHEM is a registered trademark of Merck KGaA, Darmstadt, Germany
TWEEN is a registered trademark of Croda International PLC
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Code de la classe de stockage
10 - Combustible liquids
Classe de danger pour l'eau (WGK)
WGK 1
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