MONOCLONAL ANTI- BAG-1
CLONE 3.10G3E2
Purified Mouse Immunoglobulin
Product Number B 5308
Product Description
Monoclonal Anti-BAG-1 (Bcl2 Interacting Protein; RAP
46, HAP 1) (mouse IgG1 isotype) is derived from the
hybridoma produced by the fusion of splenocytes from
BALB/c mice immunized with full length recombinant
BAG-1 protein and SP2/0 mouse myeloma cells.
Monoclonal Anti-BAG-1 recognizes three different
isoforms of human BAG-1 protein: 36 kDa
(predominant), 46 kDa (minor) and 50 kDa (minor). It
has been used in immunoblotting, immunoprecipitation,
and immunohistochemistry with frozen and formalinfixed, paraffin-embedded tissue sections.
The oncogene Bcl2 is a membrane protein that blocks a
step in a pathway leading to apoptosis. The mouse
protein that binds to Bcl2 was identified by interaction
cloning. This protein, referred to as Bcl2-Interacting
Protein-1 (BAG-1), is a heat shock 70 kDa (Hsp70)binding protein that can collaborate with Bcl-2 in
suppressing apoptosis. Overproduction of Bcl2 occurs
frequently in human cancers and contributes to tumor
radio- and chemoresistance by blocking apoptosis
induced by genotoxic injury and other types of damage.
Conversely, reduced levels of Bcl2 have been
associated with higher rates of spontaneous and
inducible apoptosis in circulating lymphocytes of
1,2
persons infected with HIV and some other viruses.
The Bcl2-interacting protein, BAG-1, enhances the antiapoptotic effects of Bcl2. BAG-1 also interacts with the
serine/threonine protein kinase Raf-1. Raf-1 and
BAG-1 proteins can also be coimmunoprecipitated from
mammalian cells and from insect cells infected with
recombinant baculoviruses encoding these proteins.
Recombinant human BAG-1 is expressed as four
isoforms, designated p50, p46, p33 and p29. The
isoforms have different subcellular localization, bind to
different proteins and play different roles in a variety of
cellular processes including signal transduction, heat
shock, apoptosis and transcription. BAG-1M (p46), one
of the isoforms of BAG-1, was reported to bind to DNA
and stimulate general transcription when cells were
3,4
stressed by heat shock.
BAG-1 represents a link between growth factor
receptors and anti-apoptotic mechanisms. BAG-1
interacts with both the hepatocyte growth factor
receptor and the platelet-derived growth factor receptor
and, in both cases, enhances growth factor-mediated
protection from apoptosis. The C-terminal region of the
BAG-1 protein was found to be responsible for binding
to the receptors, but the entire BAG-1 protein was
required for protection from apoptosis. BAG-1 is
expressed in a variety of cells in normal adult tissues
5,6
and is localized either in cytosol or nucleus, or both.
Reagent
Monoclonal Anti-BAG-1 is supplied as a solution in
phosphate buffered saline, pH 7.4, with 0.08% sodium
azide as a preservative.
Precautions and Disclaimer
Due to the sodium azide content, a material safety data
sheet (MSDS) for this product has been sent to the
attention of the safety officer of your institution. Consult
the MSDS for information regarding hazards and safe
handling practices.
Storage/Stability
Store at –20 °C. Upon initial thawing freeze the
solution in working aliquots for extended storage.
Avoid repeated freezing and thawing to prevent
denaturing the antibody. Do store in a frost-free
freezer. The antibody is stable for at least 12
months when stored appropriately. Working
dilutions should be discarded if not used within 12
hours.
Product Profile
A recommended working concentration for
immunoblotting ranges from 1 to 5 µg/ml. For
immunoprecipitation use approximately 2 µg/mg of
protein lysate. For immunohistochemical staining
a recommended working concentration of 2 to
4 µg/ml is determined by using Anti-BAG-1
antibody on formalin-fixed, paraffin-embedded
human breast carcinoma tissue. The data
demonstrate that only tissues containing BAG-1
protein stain positively, which confirms the
specificity of Anti-BAG-1 for this protein. BT474,
SKBR-3, prostate, breast and leukemia cell lines
may be used as positive controls.
Note: In order to obtain best results using different
techniques and preparations we recommend determining
optimal working concentration by titration.
References
1. Takayama, S., et al., Cloning and functional
analysis of BAG-1: a novel Bcl-2-binding protein
with anti-cell death activity. Cell, 80, 279-281
(1995).
2. Takayama, S., et al., Cloning of cDNAs encoding
the human BAG1 protein and localization of the
human BAG1 gene to chromosome 9p12.
Genomics, 35, 494-498 (1996).
3. Bimston, D., et al., BAG-1, a negative regulator of
Hsp70 chaperone activity, uncouples nucleotide
hydrolysis from substrate release., EMBO J., 17,
6871-6878 (1998).
4. Hohfeld, J., Regulation of the heat shock conjugate
Hsc70 in the mammalian cell: the characterization
of the anti–apoptotic protein BAG-1 provides novel
insights., Biol. Chem., 379, 269-274 (1998).
5. Wang, H. G., et al., Bcl-2 interacting protein, BAG1, binds to and activates the kinase Raf-1. Proc.
Natl. Acad. Sci. U S A., 93, 7063-7068 (1996).
6. Takahashi, N., et al., BAG-1M, an isoform of Bcl-2interacting protein BAG-1, enhances gene
expression driven by CMV promoter. Biochem.
Biophys. Res. Commun., 286, 807-814 (2001).
AH 11/26/01
Sigma brand products are sold through Sigma-Aldrich, Inc.
Sigma-Aldrich, Inc. warrants that its products conform to the information contained in this and other Sigma-Aldrich publications. Purchaser
must determine the suitability of the product(s) for their particular use. Additional terms and conditions may apply. Please see reverse side of
the invoice or packing slip.