Application
Western blot
Sample
Mouse Recombinant protein (C-terminus of H2A.X with full range of phosphoryla)
Gel Running Conditions
Non-reduced Non-Denaturing (Native) (Dot blot)
Loading amount
0.384 µg
Specification
C-terminus of H2A.X with full range of phosphoryla
Blocking step
Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 20°C
Other product details
Incubation time
12 hour(s) and 0 minute(s) · Temperature: 4°C · Diluent: TBST with 5% skimmed milk
Dilution
1/1000
Secondary antibody
Dilution
1/500
Name
Non-Abcam antibody was used: Bio-Rad Goat anti- rabbit Cat no. 170-6515
Host species: Goat
Clonality: Polyclonal
Conjugation: Horse Radish Peroxidase
Host species: Goat
Clonality: Polyclonal
Conjugation: Horse Radish Peroxidase
Detection
Detection method
ECL
Exposure
2 minute(s) and 30 second(s)
Additional data
Additional Notes
Figure legend:
Mouse (left):
1: SQEY (non-phosphorylated SQEY motif of mouse H2A.X peptide), 2: pSQEY (serine monophosphorylated SQEY motif of mouse H2A.X peptide), 3: SQEpY (tyrosine monophosphorylated SQEY motif of mouse H2A.X peptide), 4: pSQEpY (serine and tyrosine di-phosphorylated SQEY motif of mouse H2A.X peptide).
A: 384 ng (256 ng/uL), B: 96 ng (64 ng/uL), C: 24 ng (16 ng/uL), D: 6 (4 ng/uL), E: 1.5 (1 ng/uL)
Conclusion: No cross reactivity with other phosphorylation states observed with the analysed concentration of antibody. Dots were visualised in mouse using 1:500 antibody dilution
Human (right):
1: pSQEY (serine monophosphorylated SQEY motif of human H2A.X peptide), 2: pSQEpY (serine and tyrosine di-phosphorylated SQEY motif of human H2A.X peptide).
A: 384 ng (256 ng/uL), B: 96 ng (64 ng/uL), C: 24 ng (16 ng/uL), D: 6 (4 ng/uL), E: 1.5 (1 ng/uL)
Conclusion: No cross reactivity with other phosphorylation states observed with the analysed concentration of antibody. Dots were visualised in human using 1:1,000 antibody dilution
Abcam response
Thank you for providing your results under Abtrial program. The antibody Anti-Histone H2A.X (phospho S139) antibody [EP854(2)Y] is recognising single phospho peptide pS139 as according to your testing and per the name of the antibody.
The reviewer received a reward from Abcam’s Loyalty Program in thanks for submitting this Abreview and for helping the scientific community make better-informed decisions.
Peter Ellis
Verified customer
投稿 Jun 02 2016