All lanes : Anti-DNA Polymerase beta antibody (ab26343) at 1/500 dilution

Lane 1 : Wild-type HeLa cell lysate
Lane 2 : POLB knockout HeLa cell lysate
Lane 3 : Jurkat cell lysate

Lysates/proteins at 20 µg per lane.

Performed under reducing conditions.

Predicted band size: 38 kDa
Observed band size: 38 kDa

Lanes 1-3: Merged signal (red and green). Green - ab26343 observed at 38 kDa. Red - loading control, ab7291 observed at 50 kDa.

ab26343 Anti-DNA Polymerase beta antibody was shown to specifically react with DNA Polymerase beta in wild-type HeLa cells. Loss of signal was observed when knockout cell line ab261748 (knockout cell lysate ab257096) was used. Wild-type and DNA Polymerase beta knockout samples were subjected to SDS-PAGE. ab26343 and Anti-alpha Tubulin antibody [DM1A] - Loading Control (ab7291) were incubated overnight at 4°C at 1 in 500 dilution and 1 in 20000 dilution respectively. Blots were developed with Goat anti- Rabbit IgG H&L (IRDye^® 800CW) preadsorbed (ab216773) and Goat anti- Mouse IgG H&L (IRDye^® 680RD) preadsorbed (ab216776) secondary antibodies at 1 in 10000 dilution for 1 hour at room temperature before imaging.

 

All lanes : Anti-DNA Polymerase beta antibody (ab26343) at 0.5 µg/ml

Lane 1 : HeLa (Human epithelial carcinoma cell line) Whole Cell Lysate
Lane 2 : Jurkat whole cell lysate (ab7899)
Lane 3 : A-431 whole cell lysate (ab7909)

Lysates/proteins at 20 µg per lane.

Secondary
All lanes : Goat polyclonal to Rabbit IgG (Alexa Fluor® 680) at 1/10000 dilution

Performed under reducing conditions.

Predicted band size: 38 kDa
Observed band size: 38 kDa
Additional bands at: 150 kDa (possible cross reactivity), 48 kDa (possible cross reactivity), 90 kDa (possible cross reactivity)

Formaldehyde-fixed human small cell lung cancer tissue stained for DNA Polymerase beta using ab26343 at 1/200 dilution in immunohistochemical analysis.

See Abreview

All lanes : Anti-DNA Polymerase beta antibody (ab26343) at 1 µg/ml

Lane 1 : NIH/3T3 whole cell lysate (ab7179)
Lane 2 : MEF1 (Mouse embryonic fibroblast cell line) Whole Cell Lysate
Lane 3 : PC12 (Rat adrenal pheochromocytoma cell line) Whole Cell Lysate

Lysates/proteins at 10 µg per lane.

Secondary
All lanes : IRDye 680 Conjugated Goat Anti-Rabbit IgG (H+L) at 1/10000 dilution

Performed under reducing conditions.

Predicted band size: 38 kDa
Observed band size: 38 kDa
Additional bands at: 120 kDa (possible cross reactivity), 30 kDa (possible cross reactivity)

Image courtesy of Human Protein Atlas ab26343 staining DNA polymerase beta in Human gall bladder, showing a strong nuclear staining pattern in the columnar epithelial cells in contrast with the cells in the lamina propria. Paraffin embedded human gall bladder tissue was incubated with ab26343 (1/150 dilution) for 30 mins at room temperature. Antigen retrieval was performed by heat induction in citrate buffer pH 6.

ab26343 was tested in a tissue microarray (TMA) containing a wide range of normal and cancer tissues as well as a cell microarray consisting of a range of commonly used, well characterised human cell lines. Further results for this antibody can be found at www.proteinatlas.org

ICC/IF image of ab26343 stained T47D cells. The cells were 100% methanol fixed (5 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 ab26343 at 1µg/ml overnight at +4°C. The secondary antibody (green) was DyLight® 488 goat anti- rabbit (ab96899) 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) at a concentration of 1.43µM.