Cell Signaling Technology
XP Monoclonal Antibody

Product Pathways - Cell Cycle / Checkpoint

Phospho-Aurora A (Thr288)/Aurora B (Thr232)/Aurora C (Thr198) (D13A11) XP® Rabbit mAb #2914

Applications Reactivity Sensitivity MW (kDa) Isotype
W IF-IC F H M R Endogenous 35, 40, 48 Rabbit IgG

Applications Key:  W=Western Blotting  IF-IC=Immunofluorescence (Immunocytochemistry)  F=Flow Cytometry
Reactivity Key:  H=Human  M=Mouse  R=Rat
Species cross-reactivity is determined by western blot. Species enclosed in parentheses are predicted to react based on 100% sequence homology.

Protocols

Specificity / Sensitivity

Phospho-Aurora A (Thr288)/Aurora B (Thr232)/Aurora C (Thr198) (D13A11) XP® Rabbit mAb detects endogenous levels of Aurora A/B/C only when phosphorylated at either Thr288, Thr232 or Thr198 respectively.

Source / Purification

Monoclonal antibody is produced by immunizing animals with a synthetic peptide corresponding to residues surrounding Thr232 of human Aurora B.

Western Blotting

Western Blotting

Western blot analysis of extracts from HeLa, L929 and C6 cells, treated with 4 mM hydroxyurea for 20 hours to induce G1/S phase or treated with 100 nM paclitaxel or 100 ng/ml nocodazole for 20 hours to induce G2/M phase, using Phospho-Aurora A (Thr288)/Aurora B (Thr232)/Aurora C (Thr198) (D11A13) XP® Rabbit mAb (upper) or Aurora B/AIM1 Antibody #3094 (lower).

Flow Cytometry

Flow Cytometry

Flow cytometric analysis of untreated Jurkat cells using Phospho-Aurora A (Thr288)/Aurora B (Thr232)/Aurora C (Thr198) (D13A11) XP® Rabbit mAb compared to propidium iodide (DNA content). The boxed population indicates phospho-Aurora A (Thr288)/Aurora B (Thr232)/Aurora C (Thr198)-positive cells.

IF-IC

IF-IC

Confocal immunofluorescent analysis of HT-1080 cells using Phospho-Aurora A (Thr288)/Aurora B (Thr232)/Aurora C (Thr198) (D13A11) XP® Rabbit mAb (green), β-Tubulin (9F3) Rabbit mAb (Alexa Fluor® 555 Conjugate) #2116 (red), and Phospho-Histone H3 (Ser10) (6G3) Mouse mAb #9706 (blue).


Background

Aurora kinases belong to a highly conserved family of mitotic serine/threonine kinases with three members identified among mammals: Aurora A, B, and C (1,2). Studies on the temporal expression pattern and subcellular localization of Aurora kinases in mitotic cells suggest an association with mitotic structure. Their functional influences span from G2 phase to cytokinesis and may be involved in key cell cycle events such as centrosome duplication, chromosome bi-orientation and segregation, cleavage furrow positioning, and ingression (3). Aurora A is detected at the centrosomes, along mitotic spindle microtubules, and in the cytoplasm of mitotically proliferating cells. Aurora A protein levels are low during G1 and S phases and peak during the G2/M phase of the cell cycle. Phosphorylation of Aurora A at Thr288 in its catalytic domain increases kinase activity. Aurora A is involved in centrosome separation, maturation, and spindle assembly and stability. Expression of Aurora B protein also peaks during the G2/M phase of the cell cycle; Aurora B kinase activity peaks at the transition from metaphase to the end of mitosis. Aurora B associates with chromosomes during prophase prior to relocalizing to the spindle at anaphase. Aurora B regulates chromosome segregation through the control of microtubule-kinetochore attachment and cytokinesis. Expression of both Aurora A and Aurora B during the G2/M phase transition is tightly coordinated with histone H3 phosphorylation (4,5); overexpression of these kinases is seen in a variety of human cancers (2,4). Aurora C localizes to the centrosome from anaphase to cytokinesis and both mRNA and protein levels peak during G2/M phase. Although typical Aurora C expression is limited to the testis, overexpression of Aurora C is detected in various cancer cell lines (6).

  1. Warner, S.L. et al. (2003) Mol. Cancer Ther. 2, 589-595.
  2. Katayama , H. et al. (2003) Cancer Metastasis Rev. 22, 451-464.
  3. Andrews, P.D. et al. (2003) Curr. Opin. Cell Biol. 15, 672-683.
  4. Pascreau, G. et al. (2003) Prog. Cell Cycle Res. 5, 369-374.
  5. Crosio, C. et al. (2002) Mol. Cell. Biol. 22, 874-885.
  6. Kimura, M. et al. (1999) J. Biol. Chem. 274, 7334-7340.

Application References

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For Research Use Only. Not For Use In Diagnostic Procedures.

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