559304
Anti-SAPK/JNK Rabbit pAb
liquid, Calbiochem®
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About This Item
Kod UNSPSC:
12352203
NACRES:
NA.41
Polecane produkty
pochodzenie biologiczne
rabbit
Poziom jakości
forma przeciwciała
affinity isolated antibody
rodzaj przeciwciała
primary antibodies
klon
polyclonal
Postać
liquid
nie zawiera
preservative
reaktywność gatunkowa
mouse, rat, human
producent / nazwa handlowa
Calbiochem®
warunki przechowywania
OK to freeze
avoid repeated freeze/thaw cycles
izotyp
IgG
Warunki transportu
wet ice
temp. przechowywania
−20°C
docelowa modyfikacja potranslacyjna
unmodified
informacje o genach
human ... MAPK8(5599)
Opis ogólny
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.
Immunogen
Human
a full-length, recombinant, human p54 SAPK/JNK2 fusion protein
Zastosowanie
Immunoblotting (1:1000)
Immunocytochemistry (1:200)
Immunocytochemistry (1:200)
Ostrzeżenie
Toxicity: Standard Handling (A)
Postać fizyczna
In 150 mM NaCl, 10 mM HEPES, 50% glycerol, 0.01% BSA, pH 7.5.
Rekonstytucja
Following initial thaw, aliquot and freeze (-20°C).
Komentarz do analizy
Positive Control
UV treated HEK293 cells
UV treated HEK293 cells
Inne uwagi
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.
Informacje prawne
CALBIOCHEM is a registered trademark of Merck KGaA, Darmstadt, Germany
TWEEN is a registered trademark of Croda International PLC
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Kod klasy składowania
10 - Combustible liquids
Klasa zagrożenia wodnego (WGK)
WGK 1
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