Western blot - VDAC1 / Porin antibody (ab15895)

All lanes : Anti-VDAC1 / Porin antibody - Mitochondrial Loading Control (ab15895) at 1 µg/ml

Lane 1 : Hela Nuclear lysate
Lane 2 : Hela cell lysate
Lane 3 : A431 cell lysate
Lane 4 : Jurkat cell lysate
Lane 5 : HEK293 cell lysate
Lane 6 : Hela Nuclear lysate with VDAC1 / Porin peptide (ab16131) at 1 µg/ml
Lane 7 : Hela cell lysate with VDAC1 / Porin peptide (ab16131) at 1 µg/ml
Lane 8 : A431 cell lysate with VDAC1 / Porin peptide (ab16131) at 1 µg/ml
Lane 9 : Jurkat cell lysate with VDAC1 / Porin peptide (ab16131) at 1 µg/ml
Lane 10 : HEK293 cell lysate with VDAC1 / Porin peptide (ab16131) at 1 µg/ml

Lysates/proteins at 20 µg per lane.

Secondary
Goat polyclonal Secondary Antibody to Rabbit IgG - H&L (HRP) (ab6721) at 1/5000 dilution
developed using the ECL technique

Performed under reducing conditions.

Predicted band size : 31 kDa
Observed band size : 31 kDa

Immunocytochemistry/ Immunofluorescence - VDAC1 / Porin antibody - Mitochondrial Loading Control (ab15895)

ICC/IF image of ab15895 stained human HeLa cells. The cells were methanol fixed (5 min) and incubated with the antibody (ab15895, 5µg/ml) for 1h at room temperature. The secondary antibody (green) was Alexa Fluor® 488 goat anti-rabbit IgG (H+L) used at a 1/1000 dilution for 1h. Image-iT^TM FX Signal Enhancer was used as the primary blocking agent, 5% BSA (TBS-T) was used for all other blocking steps. DAPI was used to stain the cell nuclei (blue). Alexa Fluor® 594 phalloidin was used to label F-actin (red).

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - VDAC1 / Porin antibody - Mitochondrial Loading Control (ab15895)

Image courtesy of Human Protein Atlas

Paraffin embedded sections of human kidney were incubated with ab15895 (1/600 dilution) at room temperature for 30 mins. Antigen retrieval was performed by heat induction in citrate buffer pH 6. ab15895 was also 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 images can be found at www.proteinatlas.org

Western blot - VDAC1 / Porin antibody - Mitochondrial Loading Control (ab15895)

All lanes : Anti-VDAC1 / Porin antibody - Mitochondrial Loading Control (ab15895) at 1 µg/ml

Lane 1 : Heart (Mouse) Tissue Lysate
Lane 2 : Kidney (Mouse) Tissue Lysate
Lane 3 : Skeletal Muscle (Mouse) Tissue Lysate (ab29711)
Lane 4 : Spinal Cord (Mouse) Tissue Lysate
Lane 5 : PC12 (Rat adrenal pheochromocytoma cell line) Whole Cell Lysate
Lane 6 : Brain (Rat) Tissue Lysate - normal tissue
Lane 7 : Kidney (Rat) Whole Cell Lysate - normal tissue (ab29480)

Lysates/proteins at 10 µg per lane.

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

Performed under reducing conditions.

Predicted band size : 31 kDa
Observed band size : 31 kDa

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - VDAC1 / Porin antibody - Mitochondrial Loading Control (ab15895)

ab15895 staining mouse heart tissue section by IHC-P.  Sections were PFA fixed and subjected to heat mediated antigen retrieval in citrate buffer (pH 6) prior to blocking in 10% serum for 10 minutes at 25°C.  The primary antibody was diluted 1/600 and incubated with the sample for 1 hour at 25°C.  A biotinylated goat anti-rabbit antibody diluted 1/400 was used as the secondary.

This image is courtesy of an anonymous Abreview

Western blot - VDAC1 / Porin antibody - Mitochondrial Loading Control (ab15895)

All lanes : Anti-VDAC1 / Porin antibody - Mitochondrial Loading Control (ab15895) at 1 µg/ml

Lane 1 : Adult wild type zebrafish brain mitochondrial lysate at 20 µg
Lane 2 : Adult wild type zebrafish brain cytosolic lysate at 30 µg

Secondary
HRP-conjugated goat F(ab’)2 Fragment at 1/10000 dilution
developed using the ECL technique

Predicted band size : 31 kDa
Observed band size : 28,29 kDa (why is the actual band size different from the predicted?)


Exposure time : 1 minute

Image courtesy of an anonymous Abreview.

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - VDAC1 / Porin antibody - Mitochondrial Loading Control (ab15895)

ab15895 staining VDAC1 / Porin in human cerebellum tissue by Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections).

Tissue was fixed in paraformaldehyde and a heat mediated antigen retrieval step was performed using citrate buffer pH 6.0. Samples were then incubated with ab15895 at a 1/1000 dilution for 12 hours at 4°C. The secondary used was an undiluted HRP conjugated anti-rabbit polyclonal. Note: The big cells are purkinje cells of the cerebellum, (no nuclear staining).

Image courtesy of Dr Markus Kipp by Abreview.

Western blot - Anti-VDAC1 / Porin antibody - Mitochondrial Loading Control (ab15895)

developed using the ECL technique

Performed under reducing conditions.

Predicted band size : 31 kDa


Exposure time : 3 minutes

Western blot image using the Optiblot Reducing Electrophoresis Kit - 10 x 10 cm (4-20%) (ab119220) with the
Prism Ultra Protein Ladder (ab116028) 5µl used. We recommend using our ECL substrate kit (ab65623) .
20ug of Lysate per lane and detection using ab15895 diluted to 1ug/ml.
Lane 1: Hela cell lysate
Lane 2: Jurkat cell lysate
Lane 3: A431 cell lysate
Lane 4: HEK293 cell lysate
Lane 5: HepG2 cell lysate.

Western blot

All lanes : Anti-VDAC1 / Porin antibody - Mitochondrial Loading Control (ab15895) at 1 µg/ml

Lane 1 : Chicken liver cell lysate
Lane 2 : CHO K1 cell lysate
Lane 3 : MDCK cell lysate
Lane 4 : Chicken liver cell lysate with VDAC1 / Porin peptide (ab16131) at 1 µg/ml
Lane 5 : CHO K1 cell lysate with VDAC1 / Porin peptide (ab16131) at 1 µg/ml
Lane 6 : MDCK cell lysate with VDAC1 / Porin peptide (ab16131) at 1 µg/ml

Lysates/proteins at 20 µg per lane.

Secondary
Alexa fluor goat polyclonal anti-Rabbit IgG at 1/10000 dilution
developed using the ECL technique

Performed under reducing conditions.

Predicted band size : 31 kDa
Observed band size : 31 kDa