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Caspase-3 (human) polyclonal antibody

 
BML-SA320-0100 100 µl 329.00 USD
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Product Specification

Host:Rabbit
 
Immunogen:Recombinant human caspase-3.
 
UniProt ID:P42574
 
Source:From rabbit serum.
 
Species reactivity:Human
 
Specificity:Recognizes the proenzyme (32 kDa) and the large (17 kDa) and small (12 kDa) subunits of mature caspase-3.
 
Crossreactivity:Does not cross-react with other caspases (caspases-1, -2, -4, -10, -14).
 
Applications:WB
 
Formulation:Liquid. Neat serum containing 0.09% sodium azide.
 
Shipping:Shipped on Blue Ice
 
Long Term Storage:-20°C
 
Protocol:Western Blotting using Caspase-3 polyclonal antibody (BML-SA320) and HL60 Cell Extracts (BML-SW101, BML-SW102)

1. HL60 cell extracts were prepared from exponentially growing cells either induced for apoptosis with etoposide (BML-SW102) or not (BML-SW101). Cells were washed once with PBS, suspended at ~4x106 cells/ml in sample buffer (62.5 mM Tris/HCl, pH 6.8; 6 M urea; 10% glycerol; 2% SDS; 0.00125% bromophenol blue; 5% b-mercaptoethanol), sonicated 15 s and incubated at 65°C, 15 min.

2. Run SDS-PAGE using 20 µL of control and induced HL60 cell extract (BML-SW101, BML-SW102).

3. Transfer proteins to nitrocellulose (NC) blotting membrane.

4. Block NC with “Blotto” { 5% non-fat dry milk in TBST (50 mM Tris/HCl, pH 7.4; 150 mM NaCl; 0.1% Tween 20)} for 1 hr at RT with gentle agitation.

5.  Incubate NC with 1/2000 (recommended starting dilution factor is 1:2000 – 1:5000) of Caspase-3 polyclonal antibody (BML-SA320) in Blotto for 2 hr at RT.

6. Wash membrane with TBST x 3, 5-10 min per wash.

7. Incubate NC with secondary antibody (1/2000 in Blotto) using anti-rabbit alkaline phosphatase conjugate (ADI-SAB-301), or anti-rabbit IgG, polyclonal antibody (HRP conjugate) (ADI-SAB-300), as appropriate, for 1 hr at RT.

8. Wash membrane with TBST x 3, 5-10 min per wash.  Rinse briefly with TBS (TBST w/o the Tween).

9. Incubate with BCIP/NBT color development reagent until bands are desired darkness or develop using chemiluminescent reagents. NOTE: Color development times may vary.

These procedures are intended only as a guide.  The optimal concentrations of primary antibody and the experimental conditions must be determined by the individual user. No warranty or guarantee of performance using these procedures is made or implied.
 
Caspase-3 (human) polyclonal antibody Western blot
Western Blot Analysis of Caspase 3 (human), pAb (BML-SA320): Lane 1: MW Marker, Lane 2: Jurkat Cell Lysate, Lane 3: Jurkat + Staurosporine Cell Lysate, Lane 4: MCF-7 Cell Lysate
Caspase-3 (human) polyclonal antibody Western blot
Western blot analysis of whole cell extracts from uninduced Jurkat cells (-) and Jurkat cells induced to BS19undergo apoptosis (+) by treatment with human, recombinant soluble TRAIL [SE-721]. One hundred thousand cells were used per lane. The nitrocellulose membrane was probed with Anti-Caspase-3 [SA-320] at 1/5000. Secondary antibody was GAR-AP (1/2000) and the nitrocellulose was developed with BCIP/NBT. P and L indicate positions of the proenzyme and large subunit of the active enzyme, respectively.
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Caspase-3 (human) polyclonal antibody Western blot Caspase-3 (human) polyclonal antibody Western blot

Product Literature References

Cytochrome c Defiency Confers Apoptosome and Mitochondrial Dysfunction in African-American Men with Prostate Cancer: R. Kumar, et al.; Cancer Res. 79, 1353 (2019), Abstract;

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