Phospho-Akt (Ser473) (D9E) XP® Rabbit mAb (BSA and Azide Free)
What is a Recombinant Antibody and Why is it Important?
Recombinant antibodies offer several key advantages compared to traditional antibodies.
These include superior lot-to-lot consistency, continuous supply, and animal-free manufacturing.
As such, recombinant antibodies are seeing increased use for scientific research, especially as a means of
addressing the ongoing reproducibility crisis.
What is a Recombinant Antibody?
Traditional polyclonal and monoclonal antibodies are the product of normal B cell development and genetic recombination.
They are generated by immunizing an animal with an antigen to elicit an immune response. While polyclonal antibodies are
secreted by many different B cell clones and recognize multiple antigenic epitopes, monoclonals originate from a single B
cell clone and are specific for just one epitope.
Recombinant antibodies are monoclonal, but their production involves in vitro genetic manipulation.
After cloning the antibody genes into an expression vector, this is then transfected into an appropriate host cell line
for antibody expression. Mammalian cell lines are most commonly used for recombinant antibody production, although cell
lines of bacterial, yeast, or insect origin are also suitable.
Superior Lot-to-Lot Consistency
Because recombinant antibody production involves sequencing the antibody light and heavy chains, it is a highly controlled
and reliable process. In contrast, hybridoma-based systems for producing monoclonal antibodies are subject to genetic
drift and instability, increasing the potential for lot-to-lot variability or loss of antibody expression. Recombinant
antibodies are highly consistent from lot to lot, thereby ensuring reproducible experimental results.
In vitro methods for producing antibodies are amenable to large-scale production, meaning antibody availability is
unlikely to become a limiting factor. Moreover, since the recombinant antibody sequence is known, continuity of supply
is assured; in situations where an antibody will be used to support large, long-term studies, this can be an especially
Unlike traditional methods for antibody production, recombinant approaches avoid the need to use animals.
Where polyclonal antibodies are purified directly from the serum of the immunized host, and monoclonals are purified
from either hybridoma-derived tissue culture supernatant or ascites, recombinant antibodies are instead purified from
the tissue culture supernatants of transfected host cell lines.
Regardless of whether an antibody is polyclonal, monoclonal or recombinant, it must always be properly validated
in the intended application prior to experimental use. At CST, we adhere to the
Hallmarks of Antibody Validation™,
six complementary strategies for determining the specificity, sensitivity, and functionality of an antibody in any
given assay. By carefully tailoring these strategies to each antibody product, we guarantee that CST antibodies
will work as expected, to help you achieve results you can trust.
Phospho-Akt (Ser473) (D9E) XP® Rabbit mAb (BSA and Azide Free) #31957
This product is the carrier-free version of product #4060. All data were generated using the same antibody clone in the standard formulation which contains BSA and glycerol.
This formulation is ideal for use with technologies requiring specialized or custom antibody labeling, including fluorophores, metals, lanthanides, and oligonucleotides. It is not recommended for ChIP, ChIP-seq, CUT&RUN, or CUT&Tag assays. If you require a carrier-free formulation for chromatin profiling, please contact us. Optimal dilutions/concentrations should be determined by the end user.
Supplied in 1X PBS, BSA and Azide Free.
For standard formulation of this product see product #4060.
Store at -20°C. This product will freeze at -20°C so it is recommended to aliquot into single-use vials to avoid multiple freeze/thaw cycles. A slight precipitate may be present and can be dissolved by gently vortexing. This will not interfere with antibody performance.
Specificity / Sensitivity
Phospho-Akt (Ser473) (D9E) XP® Rabbit mAb (BSA and Azide Free) detects endogenous levels of Akt only when phosphorylated at Ser473.
Human, Mouse, Rat, Hamster, Monkey, D. melanogaster, Zebrafish, Bovine
Source / Purification
Monoclonal antibody is produced by immunizing animals with a synthetic peptide corresponding to residues surrounding Ser473 of human Akt protein.
Akt, also referred to as PKB or Rac, plays a critical role in controlling survival and apoptosis (1-3). This protein kinase is activated by insulin and various growth and survival factors to function in a wortmannin-sensitive pathway involving PI3 kinase (2,3). Akt is activated by phospholipid binding and activation loop phosphorylation at Thr308 by PDK1 (4) and by phosphorylation within the carboxy terminus at Ser473. The previously elusive PDK2 responsible for phosphorylation of Akt at Ser473 has been identified as mammalian target of rapamycin (mTOR) in a rapamycin-insensitive complex with rictor and Sin1 (5,6). Akt promotes cell survival by inhibiting apoptosis through phosphorylation and inactivation of several targets, including Bad (7), forkhead transcription factors (8), c-Raf (9), and caspase-9. PTEN phosphatase is a major negative regulator of the PI3K/Akt signaling pathway (10). LY294002 is a specific PI3 kinase inhibitor (11). Another essential Akt function is the regulation of glycogen synthesis through phosphorylation and inactivation of GSK-3α and β (12,13). Akt may also play a role in insulin stimulation of glucose transport (12). In addition to its role in survival and glycogen synthesis, Akt is involved in cell cycle regulation by preventing GSK-3β-mediated phosphorylation and degradation of cyclin D1 (14) and by negatively regulating the cyclin-dependent kinase inhibitors p27 Kip1 (15) and p21 Waf1/Cip1 (16). Akt also plays a critical role in cell growth by directly phosphorylating mTOR in a rapamycin-sensitive complex containing raptor (17). More importantly, Akt phosphorylates and inactivates tuberin (TSC2), an inhibitor of mTOR within the mTOR-raptor complex (18,19).t