Anti-Rex1 antibody

• ICC/IF

Unknown

Immunocytochemistry/ Immunofluorescence - Anti-Rex1 antibody (AB28141)

ICC/IF image of ab28141 stained mouse embryonic stem cells. The cells were 4% PFA fixed (10 min) and then incubated in 1%BSA / 10% normal goat serum / 0.3M glycine in 0.1% PBS-Tween for 1h to permeabilise the cells and block non-specific protein-protein interactions. The cells were then incubated with the antibody (ab28141, 1µg/ml) overnight at +4°C. The secondary antibody (green) was Alexa Fluor® 488 goat anti-rabbit IgG (H+L) used at a 1/1000 dilution for 1h. Alexa Fluor® 594 WGA was used to label plasma membranes (red) at a 1/200 dilution for 1h. DAPI was used to stain the cell nuclei (blue).

• WB

Unknown

Western blot - Anti-Rex1 antibody (AB28141)

In this western blot Anti-Rex1 antibody ab28141 recognizes a band at 32kDa corresponding to the endogenous Rex1 protein and a band at 38 kDa corresponding to biotinylated Rex1. The identities of the bands at 34 and 36 kDa are unknown. It is possible that they represent non-specific binding, but it may be that they represent modified forms of the non-biotinylated endogenous Rex1. The biotinylated Rex1 protein contains an N-terminal FLAG and a 25 AA peptide substrate for biotinylation. Therefore, the biotinylated protein is expected to migrate at a slightly higher molecular weight than endogenous Rex1. The large bands at the top of the image are likely to represent non-specific binding.

All lanes:

Western blot - Anti-Rex1 antibody (ab28141) at 0.5 µg/mL

Lane 1:

Lysate of 293T cells overexpressing Rex1

Lane 2:

Lysate of Mouse Embryonic Stem Cells expressing biotinylated Rex1

Lane 3:

Negative control for tagged protein - Lysate of Mouse Embryonic Stem Cells (endogenous protein present)

Lanes 4 - 7:

Lysate of various passages of Mouse Embryonic Stem Cells expressing biotinylated Rex1

Predicted band size: 35 kDa

Observed band size: 32 kDa,34 kDa,36 kDa,38 kDa

false

• WB

Project1835****

Western blot - Anti-Rex1 antibody (AB28141)

The REX1 protein in these 293T cells contains an N-terminal FLAG tag. We expect the tagged protein to migrate at a higher molecular weight than endogenous REX1.

All lanes:

Western blot - Anti-Rex1 antibody (ab28141) at 1 µg/mL

All lanes:

293T Cells Overexpressing Mouse Rex1 at 10 µg

Secondary

All lanes:

Goat polyclonal to Rabbit IgG - H&L - Pre-Adsorbed (HRP) at 1/3000 dilution

Predicted band size: 35 kDa

Observed band size: 100 kDa,40 kDa,75 kDa

false