Anti-Myc tag antibody [9E10]

• WB

Lab

Western blot - Anti-Myc tag antibody [9E10] (AB32)

False colour image of Western blot : Anti-Myc tag antibody [9E10] staining at 1/200 dilution, shown in green; Rabbit Anti-GAPDH antibody [EPR16891] (ab181602) loading control staining at 1/20000 dilution, shown in red. In Western blot, ab32 was shown to bind specifically to Myc tag. A band was observed at 57 kDa in wild-type HEK-293T cell lysates with no signal observed at this size in MYC knockout cell line ab256500 (knockout cell lysate ab263850). The band observed in the knockout lysate lane below 57 kDa is likely to represent a truncated form of Myc tag. This has not been investigated further and the functional properties of the gene product have not been determined. To generate this image, wild-type and MYC knockout HEK-293T cell lysates were analysed.First, samples were run on an SDS-PAGE gel then transferred onto a nitrocellulose membrane. Membranes were blocked in 3 % milk in TBS-0.1 % Tween^® 20 (TBS-T) before incubation with primary antibodies overnight at 4 °C. Blots were washed four times in TBS-T, incubated with secondary antibodies for 1 h at room temperature, washed again four times then imaged. Secondary antibodies used were Goat anti-Mouse IgG H&L (IRDye^® 800CW) preabsorbed (ab216772) and Goat anti-Rabbit IgG H&L (IRDye^® 680RD) preabsorbed (ab216777) at 1/20000 dilution.

All lanes:

Western blot - Anti-Myc tag antibody [9E10] (ab32) at 1/200 dilution

Lane 2:

MYC knockout HEK-293T cell lysate at 20 µg

Lane 3:

Jurkat cell lysate at 20 µg

Lane 4:

SH-SY5Y cell lysate at 20 µg

Predicted band size: 48 kDa

false

• ICC/IF

PubMed

Immunocytochemistry/ Immunofluorescence - Anti-Myc tag antibody [9E10] (AB32)

RPE1 cells grown on coverslips were transfected with βarr2-myc, grown in low serum and then fixed and stained for Kif3A (red) and ab32 (green). Insets show higher magnifications of a representative PC. Kif3A was found in the cytoplasm and at the tip of the axoneme where it was colocalized with βarr2.

Cells were incubated with primary antibodies in permeabilization buffer (PBS with 1 mg/mL bovine serum albumin (PBS-BSA) and 0.1% triton-X-100) for 45 minutes at room temperature. After two washes with PBS-BSA, cells were incubated for 30 minutes at room temperature in PBS-BSA containing secondary antibodies. After one wash with PBS-BSA and two washes in PBS, cells were laid down on microscope slides in a PBS–glycerol mix (50/50) with DAPI.

Image from Molla-Herman A et al., PLoS One. 2008;3(11):e3728. Fig 8(B).; doi: 10.1371/journal.pone.0003728. Reproduced under the Creative Commons license http://creativecommons.org/licenses/by/4.0/

• WB

Lab

Western blot - Anti-Myc tag antibody [9E10] (AB32)

Blocking and diluting buffer and concentration : 5% NFDM/TBST.

In Western blot, Anti-GAPDH antibody [EPR16891] - Loading Control (ab181602) staining at 1/1000000 dilution.

In Western blot, Anti-Myc tag antibody [9E10] - Loading Control (ab32) staining at 1/10000 dilution.

In Western blot, Anti-6X His tag® antibody [EPR20547] - Loading Control (ab213204) staining at 1/10000 dilution.

This antibody does not cross react with DR5 and DcR2.

All lanes:

Western blot - Anti-DR4 antibody [EPR28152-57] (<a href='/en-us/products/primary-antibodies/dr4-antibody-epr28152-57-ab312848'>ab312848</a>) at 1/1000 dilution

Lane 1:

293T (Human embryonic kidney epithelial cell) transfected with an empty vector (vector control), containing a myc-His- tag® whole cell lysate at 20 µg

Lane 2:

293T transfected with human DR4 expression vector containing a myc- His-tag®, whole cell lysate at 20 µg

Lane 3:

293T transfected with human DcR2 expression vector containing a myc- His-tag®, whole cell lysate at 48 µg

Lane 4:

Recombinant human DR5 fragment protein containing a His-tag®, whole cell lysate at 10 ng

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 1/100000 dilution

Observed band size: 48 kDa

false

Exposure time: 3s

• ICC/IF

AbReview61185****

Immunocytochemistry/ Immunofluorescence - Anti-Myc tag antibody [9E10] (AB32)

ab32 staining a Myc tagged protein in HeLa cells by ICC/IF (Immunocytochemistry/Immunofluorescence). Endogenous c-myc was not detected under these conditions. Cells were fixed with paraformaldehyde, permeabilized with 0.5% Triton and blocked with 5% Serum for 30 minutes at 25°C. Samples were incubated with primary antibody (1/1000 in 5% Serum) for 1 hour at 25°C. An Alexa Fluor^® 488 conjugated Goat anti-Mouse was used as a secondary antibody.

This image is courtesy of an anonymous Abreview

• ICC/IF

PubMed

Immunocytochemistry/ Immunofluorescence - Anti-Myc tag antibody [9E10] (AB32)

(A-D) Representative examples of body forming AtMORC7-MYC, AtMORC4-MYC, At-MORC1-MYC, and AtMORC6-MYC nuclei, respectively. (E) Untransformed wt nucleus subjected to the same antibody staining and imaging procedure. Left panels = anti-MYC channel; middle panels = DAPI channel (gray scaled). DAPI stains DNA, defining the position of dense chromocenters as high intensity white foci; right panels = merged channels (DAPI in blue, MYC in green). White triangles indicate examples of chromocenter adjacent AtMORC localization. Scale bars = 5 μM.

Leaves from three-week old plants were fixed in 4% paraformaldehyde in TRIS buffer (10 mM TRIS pH 7.5, 10 mM EDTA, and 100 mM NaCl) for 20 minutes and washed twice in TRIS buffer. Leaves were chopped in 200–400 microliters lysis buffer (15 mM TRIS pH 7.5, 2 mM EDTA, 0.5 mM spermine, 80 mM KCl, 20 mM NaCl, and 0.1% Triton X-100) and filtered through a 3 μM cell strainer. 5 μL of nuclei suspension was added to 12 μL of sorting buffer (100mM TRIS pH 7.5, 50mM KCl, 2mM MgCl2, 0.05% Tween-20, and 20.5% sucrose) and air dried on chloroform dipped microscope slides for two hours and then post-fixed in 4% paraformaldehyde in PBS for 20 minutes. Slides were washed three times in PBS and incubated in blocking buffer (3% BSA, and 10% horse serum in PBS) for 30 minutes at 37°C. Nuclei were incubated at 4°C overnight in mouse monoclonal antibody against c-Myc (9E10, ab32; 1/200). Slides were washed in PBS and incubated with goat anti-mouse FITC antibody (ab7064; 1/200) for 90 minutes at room temperature. Following PBS washes, nuclei were counterstained and mounted in Vectashield mounting media with DAPI. Nuclei were analyzed with a Zeiss LSM 710 Confocal microscope at 63X or 100X magnification using Zen software.

Image from Harris CJ et al., PLoS Genet. 2016;12(5):e1005998. Fig 5.; doi: 10.1371/journal.pgen.1005998. Reproduced under the Creative Commons license http://creativecommons.org/licenses/by/4.0/

• WB

Lab

Western blot - Anti-Myc tag antibody [9E10] (AB32)

False colour image of Western blot : Anti-Myc tag antibody [9E10] staining at 1/200 dilution, shown in green; Rabbit Anti-GAPDH antibody [EPR16891] (ab181602) loading control staining at 1/20000 dilution, shown in red. In Western blot, ab32 was shown to bind specifically to Myc tag. A band was observed at 57 kDa in wild-type HEK-293T cell lysates with no signal observed at this size in MYC CRISPR-Cas9 edited cell line ab256500 (CRISPR-Cas9 edited cell lysate ab263850). The band observed in the CRISPR-Cas9 edited lysate lane below 57 kDa is likely to represent a truncated form of Myc tag. This has not been investigated further and the functional properties of the gene product have not been determined. To generate this image, wild-type and MYC CRISPR-Cas9 edited HEK-293T cell lysates were analysed. First, samples were run on an SDS-PAGE gel then transferred onto a nitrocellulose membrane. Membranes were blocked in 3 % milk in TBS-0.1 % Tween^® 20 (TBS-T) before incubation with primary antibodies overnight at 4 °C. Blots were washed four times in TBS-T, incubated with secondary antibodies for 1 h at room temperature, washed again four times then imaged. Secondary antibodies used were Goat anti-Mouse IgG H&L (IRDye^® 800CW) preabsorbed (ab216772) and Goat anti-Rabbit IgG H&L (IRDye^® 680RD) preabsorbed (ab216777) at 1/20000 dilution.

Lanes 1 - 4:

Western blot - Anti-Myc tag antibody [9E10] (ab32) at 1/200 dilution

Lanes 1 - 4:

Western blot - Anti-Myc tag antibody [9E10] (<a href='/en-us/products/primary-antibodies/myc-tag-antibody-9e10-ab206486'>ab206486</a>) at 1/200 dilution

Lane 1:

Wild-type HEK-293T cell lysate at 20 µg

Lane 2:

MYC CRISPR-Cas9 edited HEK-293T cell lysate at 20 µg

Lane 2:

Western blot - Human MYC (c-Myc) knockout HEK-293T cell line (<a href='/en-us/products/cell-lines/human-myc-c-myc-knockout-hek-293t-cell-line-ab256500'>ab256500</a>)

Lane 3:

Jurkat cell lysate at 20 µg

Lane 4:

SH-SY5Y cell lysate at 20 µg

Predicted band size: 48 kDa,49 kDa

false

• WB

Project

Western blot - Anti-Myc tag antibody [9E10] (AB32)

This image was produced using the same antibody clone but different formulation, ab32.

Lysate from E. coli recombinantly expressing 11 commonly used tags including myc tag.

All lanes:

Western blot - Anti-Myc tag antibody [9E10] (ab32) at 1 µg/mL

All lanes:

Western blot - E. coli Positive Control (Escherichia coli ) Whole Cell Lysate (<a href='/en-us/products/cell-lysates/e-coli-positive-control-escherichia-coli-whole-cell-lysate-ab5395'>ab5395</a>) at 10 µg

Secondary

All lanes:

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

Predicted band size: 48 kDa,49 kDa

Observed band size: 45 kDa

true

Exposure time: 1min

• WB

Lab

Western blot - Anti-Myc tag antibody [9E10] (AB32)

Anti-SARS-Cov-2 ORF7a antibody [EPR24850-55] (ab283962) staining at 1/1000 dilution, shown in green; Mouse anti-Myc tag [9E10] (ab32) loading control staining at 1/1000 dilution, shown in magenta.

In Western blot, ab283962 was shown to bind specifically to SARS-Cov-2 ORF7a. First, samples were run on an SDS-PAGE gel then transferred onto a nitrocellulose membrane. Membranes were blocked in fluorescent western blot (TBS-based) blocking solution before incubation with primary antibodies overnight at 4 °C. Blots were washed four times in TBS-T, incubated with secondary antibodies for 1 h at room temperature, washed again four times then imaged.

Secondary antibodies used were Goat anti-Rabbit IgG H&L 800CW and Goat anti-Mouse IgG H&L 680RD at 1/20000 dilution.

All lanes:

Western blot - Anti-SARS-CoV-2 orf7a antibody [EPR24850-55] (<a href='/en-us/products/primary-antibodies/sars-cov-2-orf7a-antibody-epr24850-55-ab283962'>ab283962</a>) at 1/1000 dilution

Lane 1:

HEK-293T overexpressing Orf7a cell lysate at 40 µg

Lane 3:

HEK-293T overexpressing Orf8 cell lysate at 20 µg

Observed band size: 13 kDa

false