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9942
Pro-Apoptosis Bcl-2 Family Antibody Sampler Kit

Pro-Apoptosis Bcl-2 Family Antibody Sampler Kit #9942

Western Blotting Image 1

Western blot analysis of extracts from various cell lines using Bad (D24A9) Rabbit mAb.

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Western Blotting Image 2

Western blot analysis of extracts from COS cells, untreated or TPA-treated, using Phospho-Bad (Ser112) (40A9) Rabbit mAb (upper) or Bad Antibody #9292 (lower).

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Western Blotting Image 3

Western blot analysis of extracts from various cell lines using Bax (D2E11) Rabbit mAb. Brimmel et al. demonstated that Jurkat cells lack Bax protein expression (10).

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Western Blotting Image 4

Western blot analysis of extracts from Raji and Ramos cell lines using Bik Antibody.

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IHC-Leica® Bond™ Image 5

Immunohistochemical analysis of paraffin-embedded human colon adenocarcinoma using Bim (C34C5) Rabbit mAb performed on the Leica® Bond™ Rx.

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Western Blotting Image 6

Western blot analysis of extracts from Raji, A20 and RL-7 cells using Bim (C34C5) Rabbit mAb.

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Western Blotting Image 7

Western blot analysis of extracts from Jurkat cells, untreated or etoposide-treated (25 µM), and RS4;11 cells, untreated or okadaic acid-treated (1 µM), using BID Antibody (Human Specific).

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Western Blotting Image 8

Western blot analysis of extracts from various cell lines using Bak (D4E4) Rabbit mAb.

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Western Blotting Image 9

Western blot analysis of extracts from A549 cells, untreated (-) or treated with Doxorubicin #5927 (500 nM, overnight; +), using Puma (D30C10) Rabbit mAb (upper) or β-Actin (D6A8) Rabbit mAb #8457 (lower).

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Western Blotting Image 10

After the primary antibody is bound to the target protein, a complex with HRP-linked secondary antibody is formed. The LumiGLO® is added and emits light during enzyme catalyzed decomposition.

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IHC-P (paraffin) Image 11

Immunohistochemical analysis of paraffin-embedded human breast carcinoma, untreated (left) or lambda phosphatase treated (right), using Phospho-Bad (Ser 112) (40A9) Rabbit mAb.

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IHC-P (paraffin) Image 12

Immunohistochemical analysis of paraffin-embedded human breast caricnoma using Bax (D2E11) Rabbit mAb in the presence of control peptide (upper) or antigen-specific peptide (lower).

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Western Blotting Image 13

Western blot analysis of extracts from HeLa cells, transfected with 100 nM SignalSilence® Control siRNA (Fluorescein Conjugate) #6201 (-) or SignalSilence® Bim siRNA I #6461 or SignalSilence® Bim siRNA II (+), using Bim (C34C5) Rabbit mAb #2933 and α-Tubulin (11H10) Rabbit mAb #2125. Bim (C34C5) Rabbit mAb confirms silencing of Bim expression and α-Tubulin (11H10) Rabbit mAb is used to control for loading and specificity of Bim siRNA.

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IHC-P (paraffin) Image 14

Immunohistochemical analysis of paraffin-embedded human breast carcinoma using Bak (D4E4) Rabbit mAb.

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Western Blotting Image 15

Western blot analysis of extracts from various cell lines using Puma (D30C10) Rabbit mAb.

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IHC-P (paraffin) Image 16

Immunohistochemical analysis of paraffin embedded COS cells untreated (left) or TPA-treated (right), showing induced cytoplasmic staining using Phospho-Bad (Ser112) (40A9) Rabbit mAb.

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IHC-P (paraffin) Image 17

Immunohistochemical analysis of paraffin-embedded human lung carcinoma using Bim (C34C5) Rabbit mAb.

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IHC-P (paraffin) Image 18

Immunohistochemical analysis of paraffin-embedded human colon carcinoma using Bak (D4E4) Rabbit mAb.

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IP Image 19

Immunoprecipitation of Puma from RL-7 cell extracts using Rabbit (DA1E) mAb IgG XP® Isotype Control #3900 (lane 2) or Puma (D30C10) Rabbit mAb (lane 3). Lane 1 is 10% input. Western blot analysis was performed using Puma (D30C10) Rabbit mAb.

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IHC-P (paraffin) Image 20

Immunohistochemical analysis of paraffin-embedded human prostate carcinoma, using Phospho-Bad (Ser 112) (40A9) Rabbit Monoclonal Antibody preincubated with control peptide (left) or Phospho-Bad (Ser 112) Blocking Peptide (IHC Specific) #1026 (right).

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IHC-P (paraffin) Image 21

Immunohistochemical analysis of paraffin-embedded human lymphoma using Bim (C34C5) Rabbit mAb.

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IHC-P (paraffin) Image 22

Immunohistochemical analysis of paraffin-embedded mouse prostate using Bak (D4E4) Rabbit mAb.

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IHC-P (paraffin) Image 23

Immunohistochemical analysis of paraffin-embedded Non-Hodgkin's lymphoma, using Phospho-Bad (Ser112) (40A9) Rabbit mAb.

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IHC-P (paraffin) Image 24

Immunohistochemical analysis of paraffin-embedded human breast carcinoma using Bim (C34C5) Rabbit mAb in the presence of control peptide (left) or antigen specific peptide (right).

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Flow Cytometry Image 25

Flow cytometric analysis of SKBR3 cells using Bak (D4E4) Rabbit mAb (blue) compared to concentration-matched Rabbit (DA1E) mAb IgG XP® Isotype Control #3900 (red). Anti-rabbit IgG (H+L), F(ab')2 Fragment (Alexa Fluor® 488 Conjugate) #4412 was used as a secondary antibody.

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Flow Cytometry Image 26

Flow cytometric analysis of COS cells, untreated (blue) or TPA/Calyculin A treated (green), using Phospho-Bad (Ser112) (40A9) Rabbit mAb.

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Flow Cytometry Image 27

Flow cytometric analysis of Raji cells using Bim (C34C5) Rabbit mAb (blue) compared to a nonspecific negative control antibody (red).

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IF-IC Image 28

Confocal immunofluorescent analysis of OVCAR8 cells using Bak (D4E4) Rabbit mAb (green; left) showing colocalization with mitochondria that were labeled with MitoTracker® Red CMXRos (red; center). Blue pseudocolor = DRAQ5® #4084 (fluorescent DNA dye).

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IF-IC Image 29

Confocal immunofluorescent analysis of MCF-7 cells using Bim Antibody (green) showing colocalization with mitochondria that have been labeled with MitoTracker® Red CMXRos (red). Blue pseudocolor = DRAQ5® #4084 (fluorescent DNA dye).

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Product Includes Quantity Applications Reactivity MW(kDa) Isotype
Bad (D24A9) Rabbit mAb 9239 20 µl
  • WB
H M R Mk 23 Rabbit IgG
Phospho-Bad (Ser112) (40A9) Rabbit mAb 5284 20 µl
  • WB
  • IHC
  • F
H M R Mk 23 Rabbit IgG
Bax (D2E11) Rabbit mAb 5023 20 µl
  • WB
  • IP
  • IHC
H 20 Rabbit IgG
Bik Antibody 4592 20 µl
  • WB
H 20 Rabbit 
Bim (C34C5) Rabbit mAb 2933 20 µl
  • WB
  • IP
  • IHC
  • IF
  • F
H M R 12, 15, 23 Rabbit IgG
BID Antibody (Human Specific) 2002 20 µl
  • WB
  • IP
H 15, 22 Rabbit 
Bak (D4E4) Rabbit mAb 12105 20 µl
  • WB
  • IP
  • IHC
  • IF
  • F
H M R Mk 25 Rabbit IgG
Puma (D30C10) Rabbit mAb 12450 20 µl
  • WB
  • IP
H 23 Rabbit IgG
Anti-rabbit IgG, HRP-linked Antibody 7074 100 µl
  • WB
Goat 

The Pro-Apoptosis Bcl-2 Family Antibody Sampler Kit provides an economical means to examine several members of the Bcl-2 family and their activation status. The kit contains enough primary and secondary antibodies to perform two Western blot experiments per primary antibody.

Each antibody in the Pro- Apoptosis Bcl-2 Family Antibody Sampler Kit recognizes only its specific target. The antibodies do not cross-react with other Bcl-2 family members. Phospho-Bad (Ser112) (40A9) Rabbit mAb detects endogenous levels of Bad only when phosphorylated at Ser112 (mouse), Ser75 (human), or Ser113 (rat). Puma (D30C10) Rabbit mAb recognizes endogenous levels of total Puma protein but also cross-reacts with a protein of unknown origin at 60 kDa.

Phospho-Bad (Ser112) (40A9) Rabbit mAb is produced by immunizing animals with a synthetic phosphopeptide corresponding to residues surrounding Ser112 of mouse Bad. Total Bad, Bax, Bim, Bak and Puma monoclonal antibodies are produced by immunizing animals with synthetic peptides corresponding to residues surrounding Pro102 of human Bad, Leu45 of human Bax, Pro25 of human Bim, Gly75 of human Bak, or near the carboxy terminus of human Puma. Polyclonal antibodies are produced by immunizing animals with synthetic peptides corresponding to the amino-terminus of human Bik or residues surrounding the cleavage site of human BID. Polyclonal antibodies are purified by protein A and peptide affinity chromatography.

The Bcl-2 family consists of a number of evolutionarily conserved proteins containing Bcl-2 homology domains (BH) that regulate apoptosis through control of mitochondrial membrane permeability and release of cytochrome c (1-3). Four BH domains have been identified (BH1-4) that mediate protein interactions. The family can be separated into three groups based upon function and sequence homology: pro-survival members include Bcl-2, Bcl-xL, Mcl-1, A1 and Bcl-w; pro-apoptotic proteins include Bax, Bak and Bok; and "BH3 only" proteins Bad, Bik, Bid, Puma, Bim, Bmf, Noxa and Hrk. Interactions between death-promoting and death-suppressing Bcl-2 family members has led to a rheostat model in which the ratio of pro-apoptotic and anti-apoptotic proteins controls cell fate (4). Thus, pro-survival members exert their behavior by binding to and antagonizing death-promoting members. In general, the "BH3-only members" can bind to and antagonize the pro-survival proteins leading to increased apoptosis (5). While some redundancy of this system likely exists, tissue specificity, transcriptional and post-translational regulation of many of these family members can account for distinct physiological roles.

Bad is a pro-apoptotic member of the Bcl-2 family that can displace Bax from binding to Bcl-2 and Bcl-xL, resulting in cell death (6,7). Survival factors such as IL-3 can inhibit the apoptotic activity of Bad by activating intracellular signaling pathways that result in the phosphorylation of Bad at Ser112 and Ser136 (7). Phosphorylation at these sites results in the binding of Bad to 14-3-3 proteins and the inhibition of Bad binding to Bcl-2 and Bcl-xL (7). Akt has been shown to promote cell survival via its ability to phosphorylate Bad at Ser136 (8,9). Ser112 has been shown to be the substrate in vivo and in vitro of p90RSK (10,11) and mitochondria-anchored PKA (12).

  1. Cory, S. et al. (2003) Oncogene 22, 8590-607.
  2. Antonsson, B. and Martinou, J.C. (2000) Exp Cell Res 256, 50-7.
  3. Sharpe, J.C. et al. (2004) Biochim Biophys Acta 1644, 107-13.
  4. Korsmeyer, S.J. et al. (1993) Semin Cancer Biol 4, 327-32.
  5. Bouillet, P. and Strasser, A. (2002) J Cell Sci 115, 1567-74.
  6. Yang, E. et al. (1995) Cell 80, 285-291.
  7. Zha, J. et al. (1996) Cell 87, 619-628.
  8. Datta, S. R. et al. (1997) Cell 91, 231-241.
  9. Peso, L. et al. (1997) Science 278, 687-689.
  10. Bonni, A. et al. (1999) Science 286, 1358-1362.
  11. Tan, Y. et al. (1999) J. Bio. Chem. 274, 34859-34867.
  12. Harada, H. et al. (1999) Mol. Cell 3, 413-422.
Entrez-Gene Id
572 , 578 , 581 , 637 , 638 , 10018 , 27113
Swiss-Prot Acc.
Q92934 , Q16611 , Q07812 , P55957 , Q13323 , O43521 , Q9BXH1
For Research Use Only. Not For Use In Diagnostic Procedures.

Cell Signaling Technology is a trademark of Cell Signaling Technology, Inc.
U.S. Patent No. 7,429,487, foreign equivalents, and child patents deriving therefrom.

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To Purchase # 9942T

Product Number Size
9942T 1 Kit (8 x 20 µl)

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