추천 제품
생물학적 소스
rabbit
Quality Level
항체 형태
affinity isolated antibody
항체 생산 유형
primary antibodies
클론
polyclonal
양식
liquid
미포함
preservative
종 반응성
mouse, rat, human
제조업체/상표
Calbiochem®
저장 조건
OK to freeze
avoid repeated freeze/thaw cycles
동형
IgG
배송 상태
wet ice
저장 온도
−20°C
타겟 번역 후 변형
unmodified
유전자 정보
mouse ... Mapk11(19094)
일반 설명
면역원
애플리케이션
Immunoblotting (1:1000)
Paraffin Sections (1:50, heat pretreatment required, see comments)
경고
물리적 형태
재구성
기타 정보
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% bromophenol 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
1. Lyse cells by adding 100 ml SDS Sample Buffer and immediately scrape the cells off the plate and transfer the extract to a microfuge tube. Keep on ice.
2. Sonicate for 2 s to shear DNA and reduce sample viscosity.
3. Heat sample to 95-100°C for 5 min. Cool on ice.
4. Microcentrifuge for 5 min.
5. Load 20 ml onto SDS-PAGE gel (10 cm x 10 cm).
6. Electrotransfer to nitrocellulose membrane.
As controls, we recommend using 15 ml of phosphorylated and nonphosphorylated C-6 glioma cell extracts.
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 of 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.
Zervos, A.S., et al. 1995. Proc. Natl. Acad. Sci. USA92, 10531.
Han, J., et al. 1994. Science265, 808.
Lee, J.C., et al. 1994. Nature372, 739.
Rouse, J., et al. 1994. Cell78, 1027.
법적 정보
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Storage Class Code
10 - Combustible liquids
WGK
WGK 1
시험 성적서(COA)
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