Mouse Loading Control Antibody Panel

• WB

Unknown

Western blot - Mouse Loading Control Antibody Panel (AB199713)

Lanes 1 - 3:

Western blot - Anti-alpha 1 Sodium Potassium ATPase antibody [464.6] - BSA and Azide free (<a href='/en-us/products/primary-antibodies/alpha-1-sodium-potassium-atpase-antibody-4646-bsa-and-azide-free-ab237969'>ab237969</a>) at 1 µg/mL

Lanes 4 - 6:

Western blot - Anti-alpha 1 Sodium Potassium ATPase antibody [464.6] - BSA and Azide free (<a href='/en-us/products/primary-antibodies/alpha-1-sodium-potassium-atpase-antibody-4646-bsa-and-azide-free-ab237969'>ab237969</a>) at 2 µg/mL

Lanes 7 - 9:

Western blot - Anti-alpha 1 Sodium Potassium ATPase antibody [464.6] - BSA and Azide free (<a href='/en-us/products/primary-antibodies/alpha-1-sodium-potassium-atpase-antibody-4646-bsa-and-azide-free-ab237969'>ab237969</a>) at 5 µg/mL

Lanes 1, 4 and 7:

<a href='/en-us/products/unavailable/human-brain-tissue-lysate-total-protein-ab29466'>ab29466</a> at 20 µg

Lanes 2, 5 and 8:

ab27253 at 20 µg

Lanes 3, 6 and 9:

<a href='/en-us/products/unavailable/brain-rat-tissue-lysate-ab7942'>ab7942</a> at 20 µg

Predicted band size: 113 kDa

true

Exposure time: 3min

• WB

Unknown

Western blot - Mouse Loading Control Antibody Panel (AB199713)

This blot was produced using a 4-12% Bis-tris gel under the MOPS buffer system. The gel was run at 200V for 50 minutes before being transferred onto a Nitrocellulose membrane at 30V for 70 minutes. The membrane was then blocked for an hour using 2% Bovine Serum Albumin before being incubated with ab7291 overnight at 4°C. Antibody binding was detected using an anti-mouse antibody conjugated to HRP, and visualised using ECL development solution ab133406

All lanes:

Western blot - Anti-alpha Tubulin antibody [DM1A] - Loading Control (<a href='/en-us/products/primary-antibodies/alpha-tubulin-antibody-dm1a-loading-control-ab7291'>ab7291</a>) at 1 µg/mL

Lane 1:

HeLa (Human epithelial cell line from cervix adenocarcinoma cell line) whole cell lysate at 10 µg

Lane 2:

NIH/3T3 (Mouse embryonic fibroblast cell line) whole cell lysate at 10 µg

Lane 3:

PC12 (Rat adrenal gland pheochromocytoma cell line) whole cell lysate at 10 µg

Secondary

All lanes:

Western blot - Goat Anti-Mouse IgG H&L (HRP) preadsorbed (<a href='/en-us/products/secondary-antibodies/goat-mouse-igg-h-l-hrp-preadsorbed-ab97040'>ab97040</a>) at 1/50000 dilution

Predicted band size: 36 kDa,50 kDa

Observed band size: 50 kDa

true

Exposure time: 150s

• WB

Unknown

Western blot - Mouse Loading Control Antibody Panel (AB199713)

The membrane 1-5 was blocked in 5% Milk (1 hour). The membrane 6-10 was blocked in 5% BSA (1 hour). Abcam routinely uses 5% BSA to block however following recent customer feedback our labs investigated the effect of 5% milk blocking. We can now confirm that milk is not a suitable blocking agent and significantly decreases the signal on the membrane. We have informed all users of the antibody of this finding.

All lanes:

Western blot - Anti-GAPDH antibody [mAbcam 9484] - Loading Control (<a href='/en-us/products/primary-antibodies/gapdh-antibody-mabcam-9484-loading-control-ab9484'>ab9484</a>) at 1/1000 dilution

Lanes 1 and 6:

HeLa (Human epithelial carcinoma cell line) Nuclear Lysate at 20 µg

Lanes 2 and 7:

HeLa (Human epithelial carcinoma cell line) Whole Cell Lysate at 20 µg

Lanes 3 and 8:

Western blot - A-431 whole cell lysate (<a href='/en-us/products/cell-lysates/a-431-whole-cell-lysate-ab7909'>ab7909</a>) at 20 µg

Lanes 4 and 9:

Western blot - Jurkat whole cell lysate (<a href='/en-us/products/cell-lysates/jurkat-whole-cell-lysate-ab7899'>ab7899</a>) at 20 µg

Lanes 5 and 10:

Western blot - HEK-293 whole cell lysate (<a href='/en-us/products/cell-lysates/hek-293-whole-cell-lysate-ab7902'>ab7902</a>) at 20 µg

Secondary

All lanes:

Goat anti-Mouse (HRP conjugated) at 1/5000 dilution

Predicted band size: 36 kDa

Observed band size: 40 kDa

false

• WB

Unknown

Western blot - Mouse Loading Control Antibody Panel (AB199713)

All lanes:

Western blot - Anti-Histone H3 antibody [mAbcam 24834] - Nuclear Loading Control and ChIP Grade (<a href='/en-us/products/primary-antibodies/histone-h3-antibody-mabcam-24834-nuclear-loading-control-and-chip-grade-ab24834'>ab24834</a>) at 1 µg/mL

Lane 1:

NIH/3T3 (Mouse embryonic fibroblast cell line) Whole Cell Lysate at 10 µg

Lane 2:

Testis (Mouse) Tissue Lysate at 10 µg

Lane 3:

PC12 (Rat adrenal pheochromocytoma cell line) Whole Cell Lysate at 10 µg

Lane 4:

Testis (Rat) Tissue Lysate - normal tissue (<a href='/en-us/products/unavailable/testis-rat-tissue-lysate-normal-tissue-ab29388'>ab29388</a>) at 10 µg

Secondary

All lanes:

Western blot - Goat Anti-Mouse IgG H&L (HRP) preadsorbed (<a href='/en-us/products/secondary-antibodies/goat-mouse-igg-h-l-hrp-preadsorbed-ab97040'>ab97040</a>) at 1/5000 dilution

Predicted band size: 15 kDa

Observed band size: 18 kDa

true

Exposure time: 3min

• WB

Unknown

Western blot - Mouse Loading Control Antibody Panel (AB199713)

All lanes:

Western blot - Anti-COX IV antibody [mAbcam33985] - Mitochondrial Marker (<a href='/en-us/products/primary-antibodies/cox-iv-antibody-mabcam33985-mitochondrial-marker-ab33985'>ab33985</a>) at 1 µg/mL

Lane 1:

Jurkat (Human T cell lymphoblast-like cell line) Whole Cell Lysate at 10 µg

Lane 2:

HepG2 (Human hepatocellular liver carcinoma cell line) Whole Cell Lysate at 10 µg

Lane 3:

Human skeletal muscle tissue lysate - total protein (<a href='/en-us/products/unavailable/human-skeletal-muscle-tissue-lysate-total-protein-ab29330'>ab29330</a>) at 10 µg

Lane 4:

Skeletal Muscle (Mouse) Tissue Lysate at 10 µg

Lane 5:

Kidney (Cow) Tissue Lysate (<a href='/en-us/products/unavailable/kidney-cow-tissue-lysate-ab29073'>ab29073</a>) at 10 µg

Secondary

All lanes:

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

Predicted band size: 19 kDa

Observed band size: 15 kDa

true

Exposure time: 1min