Anti-Actin antibody - Loading Control

• IHC-P

Lab

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Actin antibody - Loading Control (AB1801)

IHC image of ab1801 staining Actin in normal human colon formalin-fixed paraffin-embedded tissue sections*, performed on a Leica Bond. The section was pre-treated using heat mediated antigen retrieval with sodium citrate buffer (pH6, epitope retrieval solution 1) for 20 mins. The section was then incubated with ab1801, 5μg/ml concentration, for 15 mins at room temperature and detected using an HRP conjugated compact polymer system. DAB was used as the chromogen. The section was then counterstained with haematoxylin and mounted with DPX. No primary antibody was used in the negative control (shown on the inset).

For other IHC staining systems (automated and non-automated) customers should optimize variable parameters such as antigen retrieval conditions, primary antibody concentration and antibody incubation times.

*Tissue obtained from the Human Research Tissue Bank, supported by the NIHR Cambridge Biomedical Research Centre

• WB

Lab

Western blot - Anti-Actin antibody - Loading Control (AB1801)

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 ab1801 overnight at 4°C. Antibody binding was detected using an anti-rabbit antibody conjugated to HRP, and visualised using ECL development solution ab133406

All lanes:

Western blot - Anti-Actin antibody - Loading Control (ab1801) at 1 µg/mL

Lane 1:

HeLa (Human epithelial carcinoma cell line) Whole Cell Lysate at 10 µg

Lane 2:

Brain (Rat) Tissue Lysate at 10 µg

Lane 3:

Brain (Mouse) Tissue Lysate at 10 µg

Lane 4:

Western blot - NIH/3T3 whole cell lysate (<a href='/en-us/products/cell-lysates/nih-3t3-whole-cell-lysate-ab7179'>ab7179</a>) at 10 µg

Secondary

All lanes:

Western blot - Goat Anti-Rabbit IgG H&L (HRP) (<a href='/en-us/products/secondary-antibodies/goat-rabbit-igg-h-l-hrp-ab97051'>ab97051</a>) at 1/50000 dilution

Predicted band size: 42 kDa

Observed band size: 42 kDa

true

Exposure time: 1min

• WB

AbReview12373****

Western blot - Anti-Actin antibody - Loading Control (AB1801)

All lanes:

Western blot - Anti-Actin antibody - Loading Control (ab1801) at 1/1000 dilution

All lanes:

Whole cell lysates prepared from HUVEC cells at 15 µg

Secondary

All lanes:

HRP-conjugated goat polyclonal to rabbit Ig at 1/5000 dilution

Predicted band size: 42 kDa

true

Exposure time: 30s

This image is courtesy of an anonymous Abreview

• WB

PubMed

Western blot - Anti-Actin antibody - Loading Control (AB1801)

Western blot analysis of HeLa cells treated for 12 hours with hesperidin (h) (2.5 μg/ml, 4,01 μM), mangiferin (5 μg/ml, 11.84 μM) (m), and hesperidin (2.5 μg/ml, 4.01 μM) in a presence of mangiferin (5 μg/ml, 11.84 μM) (h+m). Immunoblotting was performed with the following primary antibodies : Bax (ab32503), BCL2 (ab59348), beta actin (ab1801), and caspase 8. After the washing steps, the membranes were incubated with goat anti-rabbit IgG (H+L) or with goat anti-mouse IgG (H+L) HRP-conjugated secondary antibodies and detected using ECL. Densitometry was performed using Image Lab software v. 4.1 (BioRad).

Top panel : Following 12h of treatment of HeLa cells with hesperidin (h), mangiferin (m), and hesperidin in a presence of mangiferin (h+m), the mRNA levels were monitored in real - time PCR experiments. The BAX and BCL2 mRNA levels results from 2 independent experiments (n = 8) are plotted relative to glyceraldehyde-3-phosphate dehydrogenase (GAPDH) and 18S rRNA levels and expressed as a fold change over the EtOH control. Error bars represent standard derivations.

Bottom panel : Following 12h of treatment of HeLa cells with hesperidin (h), mangiferin (m), and hesperidin in a presence of mangiferin (h+m), the protein levels of Bax and BCL2 were detected with SDS-PAGE and Western Blot and related to beta actin levels.

All lanes:

Western blot - Anti-Actin antibody - Loading Control (ab1801)

Predicted band size: 42 kDa

true

Image from PLoS One. 2014; 9(3): e92128. Fig 9,•DOI: 10.1371/journal.pone.0092128 Reproduced under the Creative Commons license http://creativecommons.org/licenses/by/4.0/

• WB

Ap

Western blot - Anti-Actin antibody - Loading Control (AB1801)

All lanes:

Western blot - Anti-Actin antibody - Loading Control (ab1801) at 1 µg/mL

Lane 1:

Brain (Mouse) Tissue Lysate (ab27253) at 10 µg

Lane 2:

NIH/3T3 (Mouse) Whole Cell Lysate (ab52956) at 10 µg

Secondary

All lanes:

Western blot - Goat Anti-Rabbit IgG H&L (HRP) preadsorbed (<a href='/en-us/products/secondary-antibodies/goat-rabbit-igg-h-l-hrp-preadsorbed-ab97080'>ab97080</a>) at 1/5000 dilution

Predicted band size: 42 kDa

true

Exposure time: 30s

• WB

CiteAb

Western blot - Anti-Actin antibody - Loading Control (AB1801)

Western Blotting using Anti-Actin antibody - Loading Control, ab1801. Publication image from Liu, C. H. et al., 2016, J Biomed Sci, 26841904. Legend direct from paper.

MAO-A and MAO-B levels after amphetamine. We pretreated mice with 5ECdsAP1 aptamer before amphetamine application according to Fig. 1d; we collected tissue samples (n = 4 per group) from the VTA at 90 (60 + 30 min Fig. 1d) and 180 (60 + 30 + 90 Fig. 1d) minutes after amphetamine. We obtained protein for Western blot quantitation of MAO-A (panel a) or MAO-B (panel b) level (upper molecular fragments) using Actin (lower molecular fragments) as a reference shown in lanes 2–4 (90 min samples) or lane 5–6 (180 min). Total protein (10 µg) was used for all lanes except lanes 7 & 9. The blot was stripped and used for MAO-B after MOA-A. Lane 1 : molecular size marker of 10 KD ladder; lane 2 : 5ECdsAP1 (4 nmol/kg, icv, half dose); lanes 3 & 5 : saline (2 µl, icv); lanes 4 & 6 : 5ECdsAP1 (8 nmol/kg, icv, full dose); lanes 7, 8, 9 are controls of naive mice (no aptamer or amphetamine (with increasing amount of protein : 5, 10 and 20 µg, respectively. Because we observed no change in MAO-B level (Panel b), we determined 90 min post amphetamine is the optimal time to collect VTA samples (panel a, lane 4) for quantitative comparison of the reversal of MAO-A level in the VTA tissue from mice treated with 5EC aptamers of dsAP1, ssAP1, dsNF-kB, saline (Sal), nothing (naïve), dsSP1, dsCREB, and dsRan (panel c). Aptamer 5ECdsAP1 elevated the MAO-A by 60–100 % (t test, shown as bar graphs in panel c). N = number of mice used in the test

false

• WB

CiteAb

Western blot - Anti-Actin antibody - Loading Control (AB1801)

Western Blotting using Anti-Actin antibody - Loading Control, ab1801. Publication image from Liu, C. H. et al., 2016, J Biomed Sci, 26841904. Legend direct from paper.

MAO-A and MAO-B levels after amphetamine. We pretreated mice with 5ECdsAP1 aptamer before amphetamine application according to Fig. 1d; we collected tissue samples (n = 4 per group) from the VTA at 90 (60 + 30 min Fig. 1d) and 180 (60 + 30 + 90 Fig. 1d) minutes after amphetamine. We obtained protein for Western blot quantitation of MAO-A (panel a) or MAO-B (panel b) level (upper molecular fragments) using Actin (lower molecular fragments) as a reference shown in lanes 2–4 (90 min samples) or lane 5–6 (180 min). Total protein (10 µg) was used for all lanes except lanes 7 & 9. The blot was stripped and used for MAO-B after MOA-A. Lane 1 : molecular size marker of 10 KD ladder; lane 2 : 5ECdsAP1 (4 nmol/kg, icv, half dose); lanes 3 & 5 : saline (2 µl, icv); lanes 4 & 6 : 5ECdsAP1 (8 nmol/kg, icv, full dose); lanes 7, 8, 9 are controls of naive mice (no aptamer or amphetamine (with increasing amount of protein : 5, 10 and 20 µg, respectively. Because we observed no change in MAO-B level (Panel b), we determined 90 min post amphetamine is the optimal time to collect VTA samples (panel a, lane 4) for quantitative comparison of the reversal of MAO-A level in the VTA tissue from mice treated with 5EC aptamers of dsAP1, ssAP1, dsNF-kB, saline (Sal), nothing (naïve), dsSP1, dsCREB, and dsRan (panel c). Aptamer 5ECdsAP1 elevated the MAO-A by 60–100 % (t test, shown as bar graphs in panel c). N = number of mice used in the test

false

• WB

CiteAb

Western blot - Anti-Actin antibody - Loading Control (AB1801)

Western Blotting using Anti-Actin antibody - Loading Control, ab1801. Publication image from Liu, C. H. et al., 2016, J Biomed Sci, 26841904. Legend direct from paper.

MAO-A and MAO-B levels after amphetamine. We pretreated mice with 5ECdsAP1 aptamer before amphetamine application according to Fig. 1d; we collected tissue samples (n = 4 per group) from the VTA at 90 (60 + 30 min Fig. 1d) and 180 (60 + 30 + 90 Fig. 1d) minutes after amphetamine. We obtained protein for Western blot quantitation of MAO-A (panel a) or MAO-B (panel b) level (upper molecular fragments) using Actin (lower molecular fragments) as a reference shown in lanes 2–4 (90 min samples) or lane 5–6 (180 min). Total protein (10 µg) was used for all lanes except lanes 7 & 9. The blot was stripped and used for MAO-B after MOA-A. Lane 1 : molecular size marker of 10 KD ladder; lane 2 : 5ECdsAP1 (4 nmol/kg, icv, half dose); lanes 3 & 5 : saline (2 µl, icv); lanes 4 & 6 : 5ECdsAP1 (8 nmol/kg, icv, full dose); lanes 7, 8, 9 are controls of naive mice (no aptamer or amphetamine (with increasing amount of protein : 5, 10 and 20 µg, respectively. Because we observed no change in MAO-B level (Panel b), we determined 90 min post amphetamine is the optimal time to collect VTA samples (panel a, lane 4) for quantitative comparison of the reversal of MAO-A level in the VTA tissue from mice treated with 5EC aptamers of dsAP1, ssAP1, dsNF-kB, saline (Sal), nothing (naïve), dsSP1, dsCREB, and dsRan (panel c). Aptamer 5ECdsAP1 elevated the MAO-A by 60–100 % (t test, shown as bar graphs in panel c). N = number of mice used in the test

false