Anti-CNOT7 antibody [EPR18722] - BSA and Azide free

• IP

Supplier Data

Immunoprecipitation - Anti-CNOT7 antibody [EPR18722] - BSA and Azide free (AB240994)

CNOT7 was immunoprecipitated from 1mg of HeLa (Human epithelial cell line from cervix adenocarcinoma) whole cell lysate with ab195587 at 1/40 dilution. Western blot was performed from the immunoprecipitate using ab195587 at 1/1000 dilution. VeriBlot for IP Detection Reagent (HRP) (ab131366), was used for detection at 1/10000 dilution.

Lane 1 : HeLa whole cell lysate 10μg (Input).

Lane 2 : ab195587 IP in HeLa whole cell lysate.

Lane 3 : Rabbit monoclonal IgG (ab172730) instead of ab195587 in HeLa whole cell lysate.

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

Exposure time : 1 second.

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab195587)

All lanes:

Immunoprecipitation - Anti-CNOT7 antibody [EPR18722] (<a href='/en-us/products/primary-antibodies/cnot7-antibody-epr18722-ab195587'>ab195587</a>)

Predicted band size: 33 kDa

false

• ICC/IF

Lab

Immunocytochemistry/ Immunofluorescence - Anti-CNOT7 antibody [EPR18722] - BSA and Azide free (AB240994)

Immunofluorescent analysis of 4% paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized F9 (Mouse embryonic testicular cancer cell line) cells labeling CNOT7 with ab195587 at 1/1000 dilution, followed by Goat Anti-Rabbit IgG H&L (Alexa Fluor^® 488) (ab150077) secondary antibody at 1/1000 dilution (green). Confocal image showing cytoplasmic staining on NIH/3T3 cell line. The nuclear counter stain is DAPI (blue).

Tubulin is detected with Anti-alpha Tubulin mouse MAb (ab7291) at 1/1000 dilution, followed by Goat Anti-Mouse IgG H&L (Alexa Fluor^® 594) (ab150120) secondary antibody at 1/1000 dilution (red).
The negative controls are as follows :
-ve control 1 : ab195587at 1/1000 dilution, followed by Anti-Mouse IgG H&L (Alexa Fluor^® 594) (ab150120) secondary at 1/XXXX dilution.
-ve control 2 : Anti-alpha Tubulin mouse MAb (ab7291) at 1/1000 dilution, followed by Goat Anti-Rabbit IgG H&L (Alexa Fluor^® 488) (ab150077) secondary at 1/1000 dilution.

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab195587)

• ICC/IF

Lab

Immunocytochemistry/ Immunofluorescence - Anti-CNOT7 antibody [EPR18722] - BSA and Azide free (AB240994)

Immunofluorescent analysis of 4% paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized NIH/3T3 (Mouse embryonic fibroblast cell line) cells labeling CNOT7 with ab195587 at 1/1000 dilution, followed by Goat Anti-Rabbit IgG H&L (Alexa Fluor^® 488) (ab150077) secondary antibody at 1/1000 dilution (green). Confocal image showing cytoplasmic staining on NIH/3T3 cell line. The nuclear counter stain is DAPI (blue).

Tubulin is detected with Anti-alpha Tubulin mouse MAb (ab7291) at 1/1000 dilution, followed by Goat Anti-Mouse IgG H&L (Alexa Fluor^® 594) (ab150120) secondary antibody at 1/1000 dilution (red).
The negative controls are as follows :
-ve control 1 : ab195587at 1/1000 dilution, followed by Anti-Mouse IgG H&L (Alexa Fluor^® 594) (ab150120) secondary at 1/XXXX dilution.
-ve control 2 : Anti-alpha Tubulin mouse MAb (ab7291) at 1/1000 dilution, followed by Goat Anti-Rabbit IgG H&L (Alexa Fluor^® 488) (ab150077) secondary at 1/1000 dilution.

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab195587).

• Flow Cyt (Intra)

Lab

Flow Cytometry (Intracellular) - Anti-CNOT7 antibody [EPR18722] - BSA and Azide free (AB240994)

Intracellular Flow Cytometry analysis of 4% paraformaldehyde fixed NIH/3T3 (Mouse embryonic fibroblast cell line) cell line labelling CNOT7 with ab195587 at 1/1000 dilution (red), isotype control with ab172730 (black) and unlabelled control cells without incubation with primary antibody and secondary antibody (blue).

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab195587)

• Flow Cyt (Intra)

Supplier Data

Flow Cytometry (Intracellular) - Anti-CNOT7 antibody [EPR18722] - BSA and Azide free (AB240994)

Intracellular Flow Cytometry analysis of 4% paraformaldehyde fixed F9 (Mouse embryonic testicular cancer cell line) cells labeling CNOT7 with ab195587 at 1/1000 dilution (red), isotype control with ab172730 (black) and unlabeled control cells without incubation with primary antibody and secondary antibody (blue).

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab195587)

• WB

Lab

Western blot - Anti-CNOT7 antibody [EPR18722] - BSA and Azide free (AB240994)

This data was developed using ab195587, the same antibody clone in a different buffer formulation.

Lanes 1-3 : Merged signal (red and green). Green - ab195587 observed at 33 kDa. Red - loading control ab7291 observed at 50 kDa.

ab195587 Anti-CNOT7 antibody [EPR18722] was shown to specifically react with CNOT7 in wild-type HeLa cells. Loss of signal was observed when knockout cell line ab265811 (knockout cell lysate ab258370) was used. Wild-type and CNOT7 knockout samples were subjected to SDS-PAGE. ab195587 and Anti-alpha Tubulin antibody [DM1A] - Loading Control (ab7291) were incubated overnight at 4°C at 1 in 1000 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 20000 dilution for 1 hour at room temperature before imaging.

All lanes:

Western blot - Anti-CNOT7 antibody [EPR18722] (<a href='/en-us/products/primary-antibodies/cnot7-antibody-epr18722-ab195587'>ab195587</a>) at 1/1000 dilution

Lane 1:

Wild-type HeLa cell lysate at 20 µg

Lane 2:

CNOT7 knockout HeLa cell lysate at 20 µg

Lane 2:

Western blot - Human CNOT7 knockout HeLa cell line (<a href='/en-us/products/cell-lines/human-cnot7-knockout-hela-cell-line-ab265811'>ab265811</a>)

Lane 3:

MOLT-4 cell lysate at 20 µg

Secondary

All lanes:

Western blot - Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed (<a href='/en-us/products/secondary-antibodies/goat-rabbit-igg-h-l-irdye-800cw-preadsorbed-ab216773'>ab216773</a>) at 1/10000 dilution

Predicted band size: 33 kDa

Observed band size: 33 kDa

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