Download (C3085) - Datasheet - Sigma

Monoclonal Anti-p34cdc2
Clone A17
Mouse Ascites Fluid
Product No. C 3085
Product Description
Monoclonal Anti-p34cdc2 (mouse IgG2a isotype) is
derived from the A17 hybridoma produced by the fusion
of mouse myeloma cells and splenocytes from
immunized BALB/c mice. The C-terminal two-thirds of
Xenopus p34cdc2 expressed in E. coli was used as
immunogen.1,2
Monoclonal Anti-p34cdc2 recognizes an epitope present
in the 215-232 amino acid residues of mouse p34cdc2.
The antibody detects the p34cdc2 protein (also called
cdk1, 34 kDa) in immunoblotting,2,3
immunopreciptation,1,2 ELISA, 2 and
immunohistochemistry (frozen2 or formalin-fixed,
paraffin-embedded 3 sections). The product does not
cross-react with p32cdk2 or known cyclin-dependent
kinases. Cross-reactivity has been observed with
human,2,3 hamster, mouse, chicken and
Xenopus1 p34 cdc2. Significant improvement in the
quality and localization of staining of routinely fixed
tissues is obtained by retrieval of the antigen using a
boiling step (e.g., by microwave3) in the presence of
citrate buffer.
Cell division is a fundamental biological process,
consisting of the splitting of the cell and its genetic
material into two daughter cells. Mitosis results in the
formation of two new nuclei, each having the same
number of chromosomes as the parental nucleus.
During the cell cycle of most somatic cells, DNA
synthesis (S-phase) and mitosis (M-phase) are
separated by two "growth" stages (G1 and G2) of varying
duration. Thus, a typical eukaryotic cell sequentially
passes through G1, S, G 2, and M and back into G1
during a single cycle.4 Maturation-promoting factor
(MPF), originally found during meiosis in frog oocytes, is
a cytoplasmic factor which is highly conserved among a
wide range of species and plays a key role in the
progression of the cell cycle from interphase (G2) to
metaphase (M), in both meiosis and mitosis. One of the
components of MPF is a 34 kDa
protein with kinase activity which is encoded in the
fission yeast, Shizosaccharomyces pombe, by the
cdc2 gene (p34cdc2). The kinase activity and substrate
specificity of p34cdc2 (also known as cdk1) change
during the cell cycle. These changes have been
correlated with both the phosphorylation state of
p34cdc2 and its association with other proteins called
cyclins. Complexes of 'cyclins' and p34cdc2 play a key
role in cell cycle control. Within the complexes, the
cyclin subunit serves a regulatory role, whereas
p34cdc2 has a catalytic protein kinase activity.5 Members
of the cyclin family of proteins combine with the p34cdc2
kinase subunit to form active cdc2 kinase, which
initiates M phase of mitosis and meiosis. Deactivation
of p34cdc2 is required for exit from mitosis. Besides
involvement in the G2 to M transition, these complexes
function as key regulators of each step of the cell cycle:
p34cdc2 acts as a catalytic subunit of MPF when it forms
a complex with cyclin B.5,6 However, when p34cdc2
combines with other types of cyclins, termed G1 cyclins,
it commits the cell to DNA replication. Therefore, the
cell cycle can be considered as a p34cdc2 cycle which is
controlled by biochemical modifications such as
phosphorylation of p34cdc2 and formation of complex(es)
with other proteins, including the cyclins.7 In every
eukaryote examined, p34cdc2 contains an evolutionary
conserved 16 amino acid sequence called PSTAIR
(EGVPSTAIREISLLKE) which distinguishes p34cdc2
from other protein kinases. Nevertheless, other
cyclin-dependent kinases, like cdk2 and cdk3, contain
the PSTAIR motif. The PSTAIR region of p34cdc2 is
involved in the complex formation with cyclin B. The
availability of monoclonal antibody1,2 reacting
specifically with p34cdc2 enables the subcellular
detection and localization of p34cdc2 and examination of
substrate interactions, in a variety of organisms.
Monoclonal Anti-p34cdc2 may be used for the localization
of p34cdc2 using various immunochemical assays
including ELISA, immunoblot, immunohistochemistry
and immunoprecipitation.
Titer
A titer of 1:1,000 was determined by immunoblotting
using a COS-7 cell extract.
In order to obtain best results, it is recommended that
each user determine the optimal working dilution for
individual applications by titration assay.
Reagents
The isotype is determined using Sigma ImmunoType
Kit (Product Code ISO-1) and by a double diffusion
immunoassay using Mouse Monoclonal Antibody
Isotyping Reagents (Product Code ISO-2). The product
is provided as ascites fluid with 0.1% sodium azide as a
preservative.
Precautions and Disclaimer
Due to the sodium azide content a material safety sheet
(MSDS) for this product has been sent to the attention
of the safety officer of your institution. Consult the
MSDS for information regarding hazardous and safe
handling practices.
Storage and Stability
For continuous use, store at 2-8 °C for up to one month.
For extended storage freeze in working aliquots.
Repeated freezing and thawing is not recommended.
Storage in "frost-free" freezers is not recommended. If
slight turbidity occurs upon prolonged storage, clarify
the solution by centrifugation before use.
References
1. Kobayashi, H., et al., Molec. Biol. Cell, 3, 1279
(1992).
2. Doussis-Anagnostopoulou, I., et al., Histopathology,
24, 335 (1994).
3. Goodger, N., et al., J. Pathol., in press (1996).
4. Freeman, R., and Donoghue, D., Biochemistry, 30,
2293 (1991).
5. Yamashita, M., et al., Dev. Growth Differ., 33, 617
(1991).
6. Hirai, T., et al., Mol. Reprod. Dev., 33, 131 (1992).
7. Nurburg, C., and Nurse, P., Ann. Rev. Biochem.,
61, 441 (1992).
Pcs4/99
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