Case Study 4: eXceptional Performance of XP® Monoclonal Antibodies in All Applications
Phospho-Akt (Ser473) (D9E) XP® Rabbit mAb #4060
XMT® technology provides our scientists with access to a broad range of antibody-producing B cells unattainable with traditional technologies, allowing more comprehensive screening. Together with our rigorous validation protocols, this leads to the identification of XP® monoclonal antibodies with exceptional performance in the greatest possible range of applications. Cell Signaling Technology’s new Phospho-Akt (Ser473) (D9E) XP® Rabbit mAb #4060 is an example of an antibody with superior performance in all tested applications, previously unattainable with traditional approaches (see Table 1).
Figure 1. Western blot analysis of extracts from PC-3 cells, untreated or LY294002/wortmannin-treated, and NIH/3T3 cells, serumstarved or PDGF-treated, using Phospho-Akt (Ser473) (D9E) XP® Rabbit mAb (upper) or Akt (pan) (C67E7) Rabbit mAb #4691 (lower).
Figure 2A. Immunohistochemical analysis of paraffin-embedded human lung carcinoma using Phospho-Akt (Ser473) (D9E) XP® Rabbit mAb #4060.
Figure 2B. Immunohistochemical analysis of paraffin-embedded PTEN heterozygous mutant mouse endometrium using Phospho-Akt (Ser473) (D9E) XP® Rabbit mAb #4060. (Tissue section courtesy of Dr. Sabina Signoretti, Brigham and Women’s Hospital, Harvard Medical School, Boston, MA.).
Figure 2C. Immunohistochemical analysis using Phospho-Akt (Ser473) (D9E) XP® Rabbit mAb #4060 on SignalSlide® Phospho-Akt (Ser473) IHC Controls #8101 (paraffin-embedded LNCaP cells, untreated (left) or LY294002-treated (right)).
Figure 3. Confocal immunofluorescent analysis of C2C12 cells, LY294002-treated (left) or insulin-treated (right), using Phospho-Akt (Ser473) (D9E) XP® Rabbit mAb #4060 (green). Actin filaments have been labeled with Alexa Fluor® 555 phalloidin (red). Blue pseudocolor = DRAQ5® #4084 (fluorescent DNA dye).
Figure 4. Flow cytometric analysis of Jurkat cells, untreated (green) or treated with LY294002, wortmannin and U0126 (blue), using Phospho-Akt (Ser473) (D9E) XP® Rabbit mAb #4060 compared to a nonspecific negative control antibody (red).
| No. | Antibody | Reactivity | WB | IP | IHC | Flow | IF |
|---|---|---|---|---|---|---|---|
| 4060 | Phospho-Akt (Ser473) (D9E) XP® Rabbit mAb | H, M, R, Hm, Mk, Dm, Z, B, (C, X, Dg, Pg) | ++++ | ++++ | ++++ | ++++ | ++++ |
| 4058 | Phospho-Akt (Ser473) (193H12) Rabbit mAb | H, M, R | +++ | +++ | - | +++ | +++ |
| 3787 | Phospho-Akt (Ser473) (736E11) Rabbit mAb | H, M, (R) | - | - | ++ | - | - |
| 9271 | Phospho-Akt (Ser473) Antibody | H, M, R, Hm, Dm, B, Dg, Pg, (Mk, C, X, Hr) | +++ | ++ | - | ++ | ++ |
| 4051 | Phospho-Akt (Ser473) (587F11) Mouse mAb | H, M, R, Hm, (Mk) | +++ | +++ | - | - | - |
Table 1. Summary of Phospho-Akt (Ser473) Antibodies available from CST. Species enclosed in parentheses are predicted to react based on 100% sequence homology.
Applications Key: WB=Western Blot IP=Immunoprecipitation IHC=Immunohistochemistry Flow=Flow Cytometry IF=Immunofluorescence
Testing Data Key: (++++)=Very Highly Recommended (+++)=Highly Recommended (++)=Recommended (-)=Not Recommended N/T=Not Tested
Reactivity Key: H=Human M=Mouse R=Rat Hm=Hamster Mk=Monkey C=Chicken Dm=D.melanogaster X=Xenopus Z=Zebrafish B=Bovine Dg=Dog Pg=Pig Hr=Horse
Species enclosed in parentheses are predicted to react based on 100% sequence homology.
