Anti-GAPDH 抗体 [6C5] - Loading Control

• ICC/IF

Unknown

Immunocytochemistry/ Immunofluorescence - Anti-GAPDH antibody [6C5] - Loading Control (AB8245)

ab8245 staining GAPDH in SV40LT-SMC (Rat SV40-transfected aorta smooth cell line) cells.

The cells were fixed with 4% formaldehyde (10 minutes) permeabilized with 0.1% Triton X-100 for 5 minutes and then blocked in 1% BSA/10% normal goat serum/0.3M glycine in 0.1%PBS-Tween for 1 hour. The cells were then incubated with ab8245 at 5μg/ml and ab202272 at 1/250 dilution overnight at +4°C followed by a further incubation at room temperature for 1h with Goat Anti-Mouse IgG H&L (Alexa Fluor^® 488) preadsorbed (ab150117) (shown in green). Nuclear DNA was labeled in blue with DAPI.

Image was taken with a confocal microscope (Leica-Microsystems TCS SP8).

• ICC/IF

Unknown

Immunocytochemistry/ Immunofluorescence - Anti-GAPDH antibody [6C5] - Loading Control (AB8245)

ab8245 staining GAPDH in HeLa (Human epithelial cell line from cervix adenocarcinoma) cells.

The cells were fixed with 100% methanol (5 minutes) and then blocked in 1% BSA/10% normal goat serum/0.3M glycine in 0.1%PBS-Tween for 1 hour. The cells were then incubated with ab8245 at 5 μg/ml and ab6046 at 1 μg/ml overnight at +4°C followed by a further incubation at room temperature for 1 hour with Goat Anti-Mouse IgG H&L (Alexa Fluor^® 488) preadsorbed (ab150117) at 2 μg/ml (shown in green) and Goat Anti-Rabbit IgG H&L (Alexa Fluor^® 594) preadsorbed (ab150088) at 2 μg/ml (shown in pseudo color red). Nuclear DNA was labeled in blue with DAPI.

Negative controls : 1– Rabbit primary antibody and anti-mouse secondary antibody; 2 – Mouse primary antibody and anti-rabbit secondary antibody. Controls 1 and 2 indicate that there is no unspecific reaction between primary and secondary antibodies used.

• ICC/IF

Unknown

Immunocytochemistry/ Immunofluorescence - Anti-GAPDH antibody [6C5] - Loading Control (AB8245)

ab8245 staining GAPDH in NIH/3T3 (Mouse embryo fibroblast cell line) cells.

The cells were fixed with 4% formaldehyde (10 minutes) and then blocked in 1% BSA/10% normal goat serum/0.3M glycine in 0.1%PBS-Tween for 1 hour. The cells were then incubated with ab8245 at 1 μg/ml and ab202272 at 1/250 dilution overnight at +4°C followed by a further incubation at room temperature for 1 hour with Goat Anti-Mouse IgG H&L (Alexa Fluor^® 488) preadsorbed (ab150117) (shown in green). Nuclear DNA was labeled in blue with DAPI.

Image was taken with a confocal microscope (Leica-Microsystems TCS SP8).

• WB

Lab

Western blot - Anti-GAPDH antibody [6C5] - Loading Control (AB8245)

Western blot : Rabbit Monoclonal[EPR6959] to N WASP ab126626 staining at 1/1000 dilution, shown in green; Mouse anti GAPDH ab8245 loading control staining at 1/20000 dilution, shown in magenta. A band was observed at 62 kDa in Wild-type U-87 MG cell lysates with no signal observed at this size in WASL knockout U-87 MG cell line. To generate this image, samples were run on an SDS-PAGE gel then transferred onto a nitrocellulose membrane. Membranes were blocked in 3pc 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-Rabbit 800CW and Goat anti-Mouse 680RD at 1/20,000 dilution.

All lanes:

Western blot - Anti-N WASP antibody [EPR6959] (<a href='/products/primary-antibodies/n-wasp-antibody-epr6959-ab126626'>ab126626</a>) at 1/1000 dilution

Lane 1:

Wild-type U-87 MG at 20 µg

Lane 2:

WASL knockout U-87 MG at 20 µg

Secondary

All lanes:

Goat anti-Rabbit 800CW & Goat anti-Mouse 680RD at 1/20000 dilution

Predicted band size: 55 kDa

Observed band size: 62 kDa,36 kDa

false

• WB

Lab

Western blot - Anti-GAPDH antibody [6C5] - Loading Control (AB8245)

Lanes 1-3 : Merged signal (red and green). Green - ab133497 observed at 50 kDa. Red - loading control ab8245 observed at 36 kDa.

ab133497 Anti-Flotillin 1 antibody [EPR6041] was shown to specifically react with Flotillin 1 in wild-type HEK293T cells. Loss of signal was observed when knockout cell line ab267276 (knockout cell lysate ab257109) was used. Wild-type and Flotillin 1 knockout samples were subjected to SDS-PAGE. ab133497 and Anti-GAPDH antibody [6C5] - Loading Control (ab8245) 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-Flotillin 1 antibody [EPR6041] (<a href='/products/primary-antibodies/flotillin-1-antibody-epr6041-ab133497'>ab133497</a>) at 1/1000 dilution

Lane 1:

Wild-type HEK293T cell lysate at 20 µg

Lane 2:

Western blot - Human FLOT1 (Flotillin 1) knockout HEK-293T cell lysate (<a href='/products/cell-lysates/human-flot1-flotillin-1-knockout-hek-293t-cell-lysate-ab257109'>ab257109</a>) at 20 µg

Lane 3:

HAP1 cell lysate at 20 µg

Secondary

Lanes 1 - 3:

Western blot - Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed (<a href='/products/secondary-antibodies/goat-rabbit-igg-h-l-irdye-800cw-preadsorbed-ab216773'>ab216773</a>) at 1/20000 dilution

Lanes 1 - 3:

Western blot - Goat anti-Mouse IgG H&L (IRDye® 680RD) preadsorbed (<a href='/products/secondary-antibodies/goat-mouse-igg-h-l-irdye-680rd-preadsorbed-ab216776'>ab216776</a>) at 1/20000 dilution

Predicted band size: 47 kDa

Observed band size: 50 kDa,36 kDa

false

• WB

Lab

Western blot - Anti-GAPDH antibody [6C5] - Loading Control (AB8245)

Western blot : Rabbit Polyclonal to cGKI ab110124 staining at 1/1000 dilution, shown in green; Mouse anti GAPDH ab8245 loading control staining at 1/20000 dilution, shown in magenta. A band was observed at 76 kDa. To generate this image, samples were run on an SDS-PAGE gel then transferred onto a nitrocellulose membrane. Membranes were blocked in 3pc 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-Rabbit 800CW and Goat anti-Mouse 680RD at 1/20,000 dilution.

All lanes:

Western blot - Anti-cGKI antibody (<a href='/products/primary-antibodies/cgki-antibody-ab110124'>ab110124</a>) at 1/1000 dilution

Lane 1:

Human lung at 20 µg

Lane 2:

HL-60 at 20 µg

Secondary

All lanes:

Goat anti-Rabbit 800CW & Goat anti-Mouse 680RD at 1/20000 dilution

Predicted band size: 76 kDa

Observed band size: 76 kDa,36 kDa

false

• WB

Lab

Western blot - Anti-GAPDH antibody [6C5] - Loading Control (AB8245)

False colour image of Western blot : Anti-Lck antibody [Y123] staining at 1/1000 dilution, shown in green; Mouse anti-GAPDH antibody [6C5] (ab8245) loading control staining at 1/20000 dilution, shown in red. In Western blot, ab32149 was shown to bind specifically to Lck. A band was observed at 60 kDa in wild-type Jurkat cell lysates with no signal observed at this size in Lck knockout cell line ab273855 (knockout cell lysate ab273809). To generate this image, wild-type and Lck knockout Jurkat 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-Rabbit IgG H&L (IRDye® 800CW) preabsorbed (ab216773) and Goat anti-Mouse IgG H&L (IRDye® 680RD) preabsorbed (ab216776) at 1/20000 dilution.

All lanes:

Western blot - Anti-Lck antibody [Y123] (<a href='/products/primary-antibodies/lck-antibody-y123-ab32149'>ab32149</a>) at 1/1000 dilution

Lane 1:

Wild-type Jurkat cell lysate at 20 µg

Lane 2:

Western blot - Human LCK knockout Jurkat cell lysate (<a href='/products/cell-lysates/human-lck-knockout-jurkat-cell-lysate-ab273809'>ab273809</a>) at 20 µg

Lane 3:

Ramos cell lysate at 20 µg

Lane 4:

A549 cell lysate at 20 µg

Secondary

Lanes 1 - 4:

Western blot - Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed (<a href='/products/secondary-antibodies/goat-rabbit-igg-h-l-irdye-800cw-preadsorbed-ab216773'>ab216773</a>) at 1/20000 dilution

Lanes 1 - 4:

Western blot - Goat anti-Mouse IgG H&L (IRDye® 680RD) preadsorbed (<a href='/products/secondary-antibodies/goat-mouse-igg-h-l-irdye-680rd-preadsorbed-ab216776'>ab216776</a>) at 1/20000 dilution

Predicted band size: 58 kDa

Observed band size: 60 kDa,37 kDa

false

• WB

Lab

Western blot - Anti-GAPDH antibody [6C5] - Loading Control (AB8245)

Lanes 1 - 4 : Merged signal (red and green). Green - ab227663 observed at 160 kDa. Red - loading control ab8245 (Mouse anti-GAPDH antibody [6C5]) observed at 37kDa.

ab227663 was shown to react with CD13 in wild-type THP-1 cells in western blot with loss of signal observed in ANPEP knockout cell line ab273759 (knockout cell lysate ab275505). Wild-type and ANPEP knockout THP-1 cell lysates were subjected to SDS-PAGE. Membranes were blocked in 3% milk in TBS-T (0.1% Tween^®) before incubation with ab227663 and ab8245 (Mouse anti-GAPDH antibody [6C5]) overnight at 4°C at a 1 in 400 dilution and a 1 in 20000 dilution respectively. Blots were incubated with Goat anti-Rabbit IgG H&L (IRDye^® 800CW) preabsorbed (ab216773) and Goat anti-Mouse IgG H&L (IRDye^® 680RD) preabsorbed (ab216776) secondary antibodies at 1 in 20000 dilution for 1 hour at room temperature before imaging.

All lanes:

Western blot - Anti-CD13 antibody [SP187] (<a href='/products/primary-antibodies/cd13-antibody-sp187-ab227663'>ab227663</a>) at 1/400 dilution

Lane 1:

Wild-type THP-1 cell lysate at 30 µg

Lane 2:

Western blot - Human ANPEP (CD13) knockout THP-1 cell lysate (<a href='/products/cell-lysates/human-anpep-cd13-knockout-thp-1-cell-lysate-ab275505'>ab275505</a>) at 30 µg

Lane 3:

PANC-1 cell lysate at 30 µg

Lane 4:

HEK-293 cell lysate at 30 µg

Secondary

Lanes 1 - 4:

Western blot - Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed (<a href='/products/secondary-antibodies/goat-rabbit-igg-h-l-irdye-800cw-preadsorbed-ab216773'>ab216773</a>) at 1/20000 dilution

Lanes 1 - 4:

Western blot - Goat anti-Mouse IgG H&L (IRDye® 680RD) preadsorbed (<a href='/products/secondary-antibodies/goat-mouse-igg-h-l-irdye-680rd-preadsorbed-ab216776'>ab216776</a>) at 1/20000 dilution

Predicted band size: 109 kDa

Observed band size: 160 kDa,37 kDa

false

• WB

Lab

Western blot - Anti-GAPDH antibody [6C5] - Loading Control (AB8245)

Lanes 1-3 : Merged signal (red and green). Green - ab133574 observed at 63 kDa. Red - loading control, ab8245 observed at 37 kDa.

ab133574 Anti-nmt55 / p54nrb antibody [EPR5270] was shown to specifically react with nmt55 / p54nrb in wild-type HEK293T cells. Loss of signal was observed when knockout cell line ab266244 (knockout cell lysate ab257160) was used. Wild-type and nmt55 / p54nrb knockout samples were subjected to SDS-PAGE. ab133574 and Anti-GAPDH antibody [6C5] - Loading Control (ab8245) 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 10000 dilution for 1 hour at room temperature before imaging.

All lanes:

Western blot - Anti-nmt55 / p54nrb antibody [EPR5270] (<a href='/products/primary-antibodies/nmt55-p54nrb-antibody-epr5270-ab133574'>ab133574</a>) at 1/1000 dilution

Lane 1:

Wild-type HEK293T cell lysate at 20 µg

Lane 2:

Western blot - Human NONO (nmt55 / p54nrb) knockout HEK-293T cell lysate (<a href='/products/cell-lysates/human-nono-nmt55-p54nrb-knockout-hek-293t-cell-lysate-ab257160'>ab257160</a>) at 20 µg

Lane 3:

MOLT-4 cell lysate at 20 µg

Secondary

Lanes 1 - 3:

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

Lanes 1 - 3:

Western blot - Goat anti-Mouse IgG H&L (IRDye® 680RD) preadsorbed (<a href='/products/secondary-antibodies/goat-mouse-igg-h-l-irdye-680rd-preadsorbed-ab216776'>ab216776</a>)

Predicted band size: 54 kDa

Observed band size: 63 kDa,37 kDa

false

• WB

Lab

Western blot - Anti-GAPDH antibody [6C5] - Loading Control (AB8245)

Lane 1 : Wild-type HeLa cell lysate 20 μg

Lane 2 : WWTR1 knockout HeLa cell lysate 20 μg

Lane 3 : A549 cell lysate 20 μg

Lane 4 : K562 cell lysate 20 μg

False colour image of Western blot : Anti-TAZ antibody staining at 1 μg/ml, shown in green; Mouse anti-GAPDH antibody [6C5] (ab8245) loading control staining at 1/20000 dilution, shown in red. In Western blot, ab110239 was shown to bind specifically to TAZ. A band was observed at 52 kDa in wild-type HeLa cell lysates with no signal observed at this size in WWTR1 knockout cell line ab281598 (knockout cell lysate ab282950). To generate this image, wild-type and WWTR1 knockout HeLa cell lysates were analysed. First, samples were run on an SDS-PAGE gel then transferred onto a nitrocellulose membrane. Membranes were blocked in 5 % 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-Rabbit IgG H&L (IRDye^® 800CW) preabsorbed (ab216773) and Goat anti-Mouse IgG H&L (IRDye^® 680RD) preabsorbed (ab216776) at 1/20000 dilution.

All lanes:

Western blot - Anti-TAZ antibody (<a href='/products/primary-antibodies/taz-antibody-ab110239'>ab110239</a>) at 1 µg/mL

Lane 1:

Wild-type HeLa cell lysate at 20 µg

Lane 2:

Western blot - Human WWTR1 knockout HeLa cell lysate (ab282950) at 20 µg

Lane 3:

A549 cell lysate at 20 µg

Lane 4:

K562 cell lysate at 20 µg

Secondary

Lanes 1 - 4:

Western blot - Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed (<a href='/products/secondary-antibodies/goat-rabbit-igg-h-l-irdye-800cw-preadsorbed-ab216773'>ab216773</a>) at 1/20000 dilution

Lanes 1 - 4:

Western blot - Goat anti-Mouse IgG H&L (IRDye® 680RD) preadsorbed (<a href='/products/secondary-antibodies/goat-mouse-igg-h-l-irdye-680rd-preadsorbed-ab216776'>ab216776</a>) at 1/20000 dilution

Predicted band size: 44 kDa

Observed band size: 52 kDa,37 kDa

false

• WB

Lab

Western blot - Anti-GAPDH antibody [6C5] - Loading Control (AB8245)

Lane 1 : Wild-type HeLa cell lysate (20µg)

Lane 2 : PMS2 knockout HeLa cell lysate (20µg)

Lanes 1- 2 : Merged signal (red and green). Green - ab110638 observed at 120 kDa. Red - loading control ab8245 observed at 37 kDa.

ab110638 Anti-PMS2 antibody [EPR3947] was shown to specifically react with PMS2 in wild-type HeLa cells in western blot. Loss of signal was observed when knockout cell line ab261776 (knockout cell lysate ab257142) was used. Wild-type and PMS2 knockout samples were subjected to SDS-PAGE. Membrane was blocked for 1 hour at room temperature in 0.1% TBST with 3% non-fat dried milk. ab110638 and Anti-GAPDH antibody [6C5] - Loading Control (ab8245) 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-PMS2 antibody [EPR3947] (<a href='/products/primary-antibodies/pms2-antibody-epr3947-ab110638'>ab110638</a>) at 1/1000 dilution

Lane 1:

Wild-type HeLa cell lysate at 20 µg

Lane 2:

Western blot - Human PMS2 knockout HeLa cell lysate (<a href='/products/cell-lysates/human-pms2-knockout-hela-cell-lysate-ab257142'>ab257142</a>) at 20 µg

Secondary

Lanes 1 - 2:

Western blot - Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed (<a href='/products/secondary-antibodies/goat-rabbit-igg-h-l-irdye-800cw-preadsorbed-ab216773'>ab216773</a>) at 1/20000 dilution

Lanes 1 - 2:

Western blot - Goat anti-Mouse IgG H&L (IRDye® 680RD) preadsorbed (<a href='/products/secondary-antibodies/goat-mouse-igg-h-l-irdye-680rd-preadsorbed-ab216776'>ab216776</a>) at 1/20000 dilution

Predicted band size: 96 kDa

Observed band size: 120 kDa,37 kDa

false

• WB

Lab

Western blot - Anti-GAPDH antibody [6C5] - Loading Control (AB8245)

Lanes 1- 2 : Merged signal (red and green). Green - ab124923 observed at 75 kDa. Red - Anti-GAPDH antibody [6C5] - Loading Control (ab8245) observed at 37 kDa.

ab124923 was shown to react with TLS/FUS in wild-type HEK-293T cells in western blot. Loss of signal was observed when knockout cell line ab266587 (knockout cell lysate ab257100) was used. Wild-type HEK-293T and FUS knockout HEK-293T cell lysates were subjected to SDS-PAGE. Membrane was blocked for 1 hour at room temperature in 0.1% TBST with 3% non-fat dried milk. ab124923 and Anti-GAPDH antibody [6C5] - Loading Control (ab8245) overnight at 4°C at a 1 in 1000 dilution and a 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-TLS/FUS antibody [EPR5812] (<a href='/products/primary-antibodies/tls-fus-antibody-epr5812-ab124923'>ab124923</a>) at 1/1000 dilution

Lane 1:

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

Lane 2:

Western blot - Human FUS (TLS) knockout HEK-293T cell lysate (<a href='/products/cell-lysates/human-fus-tls-knockout-hek-293t-cell-lysate-ab257100'>ab257100</a>) at 20 µg

Secondary

Lanes 1 - 2:

Western blot - Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed (<a href='/products/secondary-antibodies/goat-rabbit-igg-h-l-irdye-800cw-preadsorbed-ab216773'>ab216773</a>) at 1/20000 dilution

Lanes 1 - 2:

Western blot - Goat anti-Mouse IgG H&L (IRDye® 680RD) preadsorbed (<a href='/products/secondary-antibodies/goat-mouse-igg-h-l-irdye-680rd-preadsorbed-ab216776'>ab216776</a>) at 1/20000 dilution

Predicted band size: 53 kDa

Observed band size: 75 kDa,37 kDa

false

• WB

Lab

Western blot - Anti-GAPDH antibody [6C5] - Loading Control (AB8245)

False colour image of Western blot : Anti-OXCT1/SCOT antibody staining at 0.04 μg/ml, shown in green; Mouse anti-GAPDH antibody [6C5] (ab8245) loading control staining at 1/20000 dilution, shown in red. In Western blot, ab224250 was shown to bind specifically to OXCT1/SCOT. A band was observed at 56 kDa in wild-type HeLa cell lysates with no signal observed at this size in OXCT1 knockout cell line ab265358 (knockout cell lysate ab258557). To generate this image, wild-type and OXCT1 knockout HeLa 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-Rabbit IgG H&L (IRDye^® 800CW) preabsorbed (ab216773) and Goat anti-Mouse IgG H&L (IRDye^® 680RD) preabsorbed (ab216776) at 1/20000 dilution.

All lanes:

Western blot - Anti-OXCT1/SCOT antibody (<a href='/products/primary-antibodies/oxct1-scot-antibody-ab224250'>ab224250</a>) at 0.04 µg/mL

Lane 1:

Wild-type HeLa cell lysate at 18 µg

Lane 2:

Western blot - Human OXCT1 (SCOT) knockout HeLa cell lysate (<a href='/products/cell-lysates/human-oxct1-scot-knockout-hela-cell-lysate-ab258557'>ab258557</a>) at 18 µg

Lane 3:

Jurkat cell lysate at 18 µg

Lane 4:

A549 cell lysate at 18 µg

Secondary

Lanes 1 - 4:

Western blot - Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed (<a href='/products/secondary-antibodies/goat-rabbit-igg-h-l-irdye-800cw-preadsorbed-ab216773'>ab216773</a>) at 1/20000 dilution

Lanes 1 - 4:

Western blot - Goat anti-Mouse IgG H&L (IRDye® 680RD) preadsorbed (<a href='/products/secondary-antibodies/goat-mouse-igg-h-l-irdye-680rd-preadsorbed-ab216776'>ab216776</a>) at 1/20000 dilution

Predicted band size: 56 kDa

Observed band size: 56 kDa,37 kDa

false

• WB

Lab

Western blot - Anti-GAPDH antibody [6C5] - Loading Control (AB8245)

Lane 1 : Wild-type HeLa cell lysate (20µg)

Lane 2 : IDH1 knockout HeLa cell lysate (20µg)

Lanes 1- 2 : Merged signal (red and green). Green - ab230949 observed at 46 kDa. Red - loading control ab8245 observed at 37 kDa.

ab230949 Anti-IDH1 antibody [EPR21002] was shown to specifically react with IDH1 in wild-type HeLa cells in western blot. Loss of signal was observed when knockout cell line ab264916 (knockout cell lysate ab257221) was used. Wild-type and IDH1 knockout samples were subjected to SDS-PAGE. Membrane was blocked for 1 hour at room temperature in 0.1% TBST with 3% non-fat dried milk. ab230949 and Anti-GAPDH antibody [6C5] - Loading Control (ab8245) 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-IDH1 antibody [EPR21002] (<a href='/products/primary-antibodies/idh1-antibody-epr21002-ab230949'>ab230949</a>) at 1/1000 dilution

Lane 1:

Wild-type HeLa cell lysate at 20 µg

Lane 2:

Western blot - Human IDH1 knockout HeLa cell lysate (<a href='/products/cell-lysates/human-idh1-knockout-hela-cell-lysate-ab257221'>ab257221</a>) at 20 µg

Secondary

Lanes 1 - 2:

Western blot - Goat anti-Mouse IgG H&L (IRDye® 680RD) preadsorbed (<a href='/products/secondary-antibodies/goat-mouse-igg-h-l-irdye-680rd-preadsorbed-ab216776'>ab216776</a>) at 1/20000 dilution

Lanes 1 - 2:

Western blot - Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed (<a href='/products/secondary-antibodies/goat-rabbit-igg-h-l-irdye-800cw-preadsorbed-ab216773'>ab216773</a>) at 1/20000 dilution

Predicted band size: 46 kDa

Observed band size: 46 kDa,37 kDa

false

• WB

Lab

Western blot - Anti-GAPDH antibody [6C5] - Loading Control (AB8245)

Lanes 1 - 4 : Merged signal (red and green). Green - ab119695 observed at 52 kDa. Red - loading control, ab8245 (Mouse anti-GAPDH antibody [6C5]) observed at 37kDa.

ab119695 was shown to react with KRT14 in A431 wild-type cells in Western blot. Loss of signal was observed when KRT14 knockout sample was used. A431 wild-type and KRT14 knockout cell lysates were subjected to SDS-PAGE. Membranes were blocked in 3% Milk in TBS-T (0.1% Tween^®) before incubation with ab119695 and ab8245 (Mouse anti-GAPDH antibody [6C5]) overnight at 4°C at a 1 in 93 dilution and a 1 in 20000 dilution respectively. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye^® 800CW) preabsorbed (ab216773) and Goat anti-Mouse IgG H&L (IRDye^® 680RD) preabsorbed (ab216776) secondary antibodies at 1 in 20000 dilution for 1 hour at room temperature before imaging.

All lanes:

Western blot - Anti-Cytokeratin 14 antibody [SP53] (<a href='/products/primary-antibodies/cytokeratin-14-antibody-sp53-ab119695'>ab119695</a>) at 1/93 dilution

Lane 1:

Wild-type A-431 (Human epidermoid carcinoma cell line) whole cell lysate at 20 µg

Lane 2:

Western blot - Human KRT14 (Cytokeratin 14) knockout A-431 cell lysate (<a href='/products/cell-lysates/human-krt14-cytokeratin-14-knockout-a-431-cell-lysate-ab261706'>ab261706</a>) at 20 µg

Lane 3:

Human skin whole tissue lysate at 20 µg

Lane 4:

MCF7 (Human breast adenocarcinoma cell line) whole cell lysate at 20 µg

Secondary

Lanes 1 - 4:

Western blot - Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed (<a href='/products/secondary-antibodies/goat-rabbit-igg-h-l-irdye-800cw-preadsorbed-ab216773'>ab216773</a>) at 1/20000 dilution

Lanes 1 - 4:

Western blot - Goat anti-Mouse IgG H&L (IRDye® 680RD) preadsorbed (<a href='/products/secondary-antibodies/goat-mouse-igg-h-l-irdye-680rd-preadsorbed-ab216776'>ab216776</a>) at 1/20000 dilution

Predicted band size: 52 kDa

Observed band size: 52 kDa,37 kDa

false

• WB

Lab

Western blot - Anti-GAPDH antibody [6C5] - Loading Control (AB8245)

Western blot : Anti-TRPA1/TSA antibody [EPR26211-139] ab320715 staining at 1/1000 dilution, shown in green; Mouse anti GAPDH ab8245 loading control staining at 1/20000 dilution, shown in magenta. A band was observed at 128, 150 kDa in Wild-type A549 cell lysates with no signal observed at this size in TRPA1 knockout A549 cell line. To generate this image, samples were run on an SDS-PAGE gel then transferred onto a nitrocellulose membrane. Membranes were blocked in 3pc 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-Rabbit 800CW and Goat anti-Mouse 680RD at 1/20,000 dilution.

All lanes:

Western blot - Anti-TRPA1/TSA antibody [EPR26211-139] (<a href='/products/primary-antibodies/trpa1-tsa-antibody-epr26211-139-ab320715'>ab320715</a>) at 1/1000 dilution

Lane 1:

Wild-type A549 at 20 µg

Lane 2:

Western blot - Human TRPA1 Knockout A549 cell line (ab314995) at 20 µg

Lane 3:

U-2 OS at 20 µg

Lane 4:

PC-3 at 20 µg

Secondary

All lanes:

Goat anti-Rabbit 800CW & Goat anti-Mouse 680RD at 1/20000 dilution

Predicted band size: 128 kDa

Observed band size: 128 kDa,150 kDa,37 kDa

false

• WB

Lab

Western blot - Anti-GAPDH antibody [6C5] - Loading Control (AB8245)

Western blot : Anti-Integrin beta 1 antibody [EPR16896] ab179472 staining at 1/1000 dilution, shown in green; Mouse anti GAPDH ab8245 loading control staining at 1/20000 dilution, shown in magenta. A band was observed at 100-130 kDa in Wild-type A549 cell lysates with no signal observed at this size in ITGB1 knockout A549 cell line. To generate this image, samples were run on an SDS-PAGE gel then transferred onto a nitrocellulose membrane. Membranes were blocked in 5pc BSA 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-Rabbit 800CW and Goat anti-Mouse 680RD at 1/20,000 dilution.

All lanes:

Western blot - Anti-Integrin beta 1 antibody [EPR16896] (<a href='/products/primary-antibodies/integrin-beta-1-antibody-epr16896-ab179472'>ab179472</a>) at 1/1000 dilution

Lane 1:

Wild-type A549 at 20 µg

Lane 2:

Western blot - Human ITGB1 knockout A549 cell line (<a href='/products/cell-lines/human-itgb1-knockout-a549-cell-line-ab286593'>ab286593</a>) at 20 µg

Lane 3:

Wild-type HCT 116 at 20 µg

Lane 4:

ITGB1 knockout HCT 116 cell line at 20 µg

Secondary

All lanes:

Goat anti-Rabbit 800CW & Goat anti-Mouse 680RD at 1/20000 dilution

Predicted band size: 88 kDa

Observed band size: 100-130 kDa,37 kDa

false

• WB

Lab

Western blot - Anti-GAPDH antibody [6C5] - Loading Control (AB8245)

Lane 1 : Wild-type HeLa cell lysate (20 μg)

Lane 2 : EIF4EBP1 knockout HeLa cell lysate (20 μg)

Lanes 1-2 : Merged signal (red and green). Green - ab32024 observed at 13 kDa. Red - loading control ab8245 observed at 37 kDa.

ab32024 Anti-eIF4EBP1 antibody [Y329] was shown to specifically react with eIF4EBP1 in wild-type HeLa cells. Loss of signal was observed when knockout cell line ab264784 (knockout cell lysate ab257146) was used. Wild-type and eIF4EBP1 knockout samples were subjected to SDS-PAGE. ab32024 and Anti-GAPDH antibody [6C5] - Loading Control (ab8245) were incubated overnight at 4°C at 1 in 5000 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-eIF4EBP1 antibody [Y329] (<a href='/products/primary-antibodies/eif4ebp1-antibody-y329-ab32024'>ab32024</a>) at 1/5000 dilution

Lane 1:

Wild-type HeLa cell lysate at 20 µg

Lane 2:

Western blot - Human EIF4EBP1 knockout HeLa cell lysate (<a href='/products/cell-lysates/human-eif4ebp1-knockout-hela-cell-lysate-ab257146'>ab257146</a>) at 20 µg

Secondary

Lanes 1 - 2:

Western blot - Goat anti-Mouse IgG H&L (IRDye® 680RD) preadsorbed (<a href='/products/secondary-antibodies/goat-mouse-igg-h-l-irdye-680rd-preadsorbed-ab216776'>ab216776</a>) at 1/20000 dilution

Lanes 1 - 3:

Western blot - Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed (<a href='/products/secondary-antibodies/goat-rabbit-igg-h-l-irdye-800cw-preadsorbed-ab216773'>ab216773</a>) at 1/20000 dilution

Predicted band size: 13 kDa

Observed band size: 13 kDa,37 kDa

false

• WB

Lab

Western blot - Anti-GAPDH antibody [6C5] - Loading Control (AB8245)

Lanes 1-3 : Merged signal (red and green). Green - ab134937 observed at 21 kDa. Red - loading control ab8245 observed at 36 kDa.

ab134937 Anti-ITPA antibody [EPR8780] was shown to specifically react with ITPA in wild-type HEK293T cells. Loss of signal was observed when knockout cell line ab266679 (knockout cell lysate ab258477) was used. Wild-type and ITPA knockout samples were subjected to SDS-PAGE. ab134937 and Anti-GAPDH antibody [6C5] - Loading Control (ab8245) 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-ITPA antibody [EPR8780] (<a href='/products/primary-antibodies/itpa-antibody-epr8780-ab134937'>ab134937</a>) at 1/1000 dilution

Lane 1:

Wild-type HEK293T cell lysate at 20 µg

Lane 2:

Western blot - Human ITPA knockout HEK-293T cell lysate (<a href='/products/cell-lysates/human-itpa-knockout-hek-293t-cell-lysate-ab258477'>ab258477</a>) at 20 µg

Lane 3:

Jurkat cell lysate at 20 µg

Secondary

Lanes 1 - 3:

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

Lanes 1 - 3:

Western blot - Goat anti-Mouse IgG H&L (IRDye® 680RD) preadsorbed (<a href='/products/secondary-antibodies/goat-mouse-igg-h-l-irdye-680rd-preadsorbed-ab216776'>ab216776</a>) at 1/20000 dilution

Predicted band size: 21 kDa

Observed band size: 36 kDa,21 kDa

false

• WB

Lab

Western blot - Anti-GAPDH antibody [6C5] - Loading Control (AB8245)

Western blot : Anti-FMRP antibody ab109741 staining at 1 µg/mL, shown in green; Mouse anti GAPDH (ab8245) loading control staining at 1/20,000 dilution, shown in magenta. A band was observed at 75 kDa in Wild-type U-87 MG cell lysates with no signal observed at this size in FMR1 knockout U-87 MG cell line. To generate this image, samples were run on an SDS-PAGE gel then transferred onto a nitrocellulose membrane. Membranes were blocked in 5 % 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 Donkey anti-Goat 800CW & Donkey anti-Mouse 680RD at 1/20,000 dilution.

All lanes:

Western blot - Anti-FMRP antibody (<a href='/products/primary-antibodies/fmrp-antibody-ab109741'>ab109741</a>) at 1 µg/mL

Lane 1:

Wild-type U-87 MG at 20 µg

Lane 2:

Western blot - Human FMR1 knockout U-87 MG cell line (<a href='/products/cell-lines/human-fmr1-knockout-u-87-mg-cell-line-ab306664'>ab306664</a>) at 20 µg

Lane 3:

Wild-type A549 at 20 µg

Lane 4:

Western blot - Human FMR1 knockout A549 cell line (<a href='/products/cell-lines/human-fmr1-knockout-a549-cell-line-ab288956'>ab288956</a>) at 20 µg

Secondary

All lanes:

Donkey anti-Goat 800CW & Donkey anti-Mouse 680RD at 1/20000 dilution

Predicted band size: 71 kDa

Observed band size: 75 kDa,36 kDa

false

• WB

Lab

Western blot - Anti-GAPDH antibody [6C5] - Loading Control (AB8245)

Western blot : Rabbit Monoclonal[EPR23994-103] to Calcium-independent Phospholipase A2/PLA2G6 ab259950 staining at 1/1000 dilution, shown in green; Mouse anti GAPDH (ab8245) loading control staining at 1/20,000 dilution, shown in magenta. A band was observed at 76 kDa in Wild-type U-87 MG cell lysates with no signal observed at this size in PLA2G6 knockout U-87 MG cell line. To generate this image, 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-Rabbit 800CW & Goat anti-Mouse 680RD at 1/20,000 dilution.

All lanes:

Western blot - Anti-Calcium-independent Phospholipase A2/PLA2G6 antibody [EPR23994-103] (<a href='/products/primary-antibodies/calcium-independent-phospholipase-a2-pla2g6-antibody-epr23994-103-ab259950'>ab259950</a>) at 1/1000 dilution

Lane 1:

Wild-type U-87 MG at 20 µg

Lane 2:

Western blot - Human PLA2G6 knockout U-87 MG cell line (<a href='/products/cell-lines/human-pla2g6-knockout-u-87-mg-cell-line-ab306752'>ab306752</a>) at 20 µg

Lane 3:

LNCaP at 20 µg

Lane 4:

Human Brain at 20 µg

Lane 5:

PC-3 at 20 µg

Secondary

All lanes:

Goat anti-Rabbit 800CW & Goat anti-Mouse 680RD at 1/20000 dilution

Predicted band size: 90 kDa

Observed band size: 76 kDa

false

• WB

Lab

Western blot - Anti-GAPDH antibody [6C5] - Loading Control (AB8245)

Western blot : Anti-FMRP antibody ab17722 staining at 1 µg/mL, shown in green; Mouse anti GAPDH (ab8245) loading control staining at 1/20,000 dilution, shown in magenta. A band was observed at 75 kDa in Wild-type U-87 MG cell lysates with no signal observed at this size in FMR1 knockout U-87 MG cell line. To generate this image, samples were run on an SDS-PAGE gel then transferred onto a nitrocellulose membrane. Membranes were blocked in 5 % 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-Rabbit 800CW & Goat anti-Mouse 680RD at 1/20,000 dilution.

All lanes:

Western blot - Anti-FMRP antibody (<a href='/products/primary-antibodies/fmrp-antibody-ab17722'>ab17722</a>) at 1 µg/mL

Lane 1:

Wild-type U-87 MG at 20 µg

Lane 2:

Western blot - Human FMR1 knockout U-87 MG cell line (<a href='/products/cell-lines/human-fmr1-knockout-u-87-mg-cell-line-ab306664'>ab306664</a>) at 20 µg

Lane 3:

Wild-type A549 at 20 µg

Lane 4:

Western blot - Human FMR1 knockout A549 cell line (<a href='/products/cell-lines/human-fmr1-knockout-a549-cell-line-ab288956'>ab288956</a>) at 20 µg

Secondary

All lanes:

Goat anti-Rabbit 800CW & Goat anti-Mouse 680RD at 1/20000 dilution

Predicted band size: 71 kDa

Observed band size: 75 kDa,36 kDa

false

• WB

Lab

Western blot - Anti-GAPDH antibody [6C5] - Loading Control (AB8245)

Western blot : Rabbit Polyclonal to MeCP2 ab195393 staining at 1/1000 dilution, shown in green; Mouse anti GAPDH (ab8245) loading control staining at 1/20,000 dilution, shown in magenta. A band was observed at 70 kDa in Wild-type U-87 MG cell lysates with no signal observed at this size in MECP2 knockout U-87 MG cell line. To generate this image, 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-Rabbit 800CW & Goat anti-Mouse 680RD at 1/20,000 dilution.

All lanes:

Western blot - Anti-MeCP2 antibody - ChIP Grade (<a href='/products/primary-antibodies/mecp2-antibody-chip-grade-ab195393'>ab195393</a>) at 1/1000 dilution

Lane 1:

Wild-type U-87 MG at 20 µg

Lane 2:

Western blot - Human MECP2 knockout U-87 MG cell line (ab306725) at 20 µg

Lane 3:

Wild-type HAP1 at 20 µg

Lane 4:

MECP2 knockout HAP1 at 20 µg

Secondary

All lanes:

Goat anti-Rabbit 800CW & Goat anti-Mouse 680RD at 1/20000 dilution

Predicted band size: 52 kDa

Observed band size: 70 kDa,36 kDa

false

• WB

Lab

Western blot - Anti-GAPDH antibody [6C5] - Loading Control (AB8245)

Lanes 1-4 : Merged signal (red and green). Green - ab191393 observed at 20 kDa. Red - loading control ab8245 observed at 37 kDa.

ab191393 Anti-MYL9 antibody [EPR13012(2)] was shown to specifically react with MYL9 in wild-type HeLa cells. Loss of signal was observed when knockout cell line ab266036 (knockout cell lysate ab256999) was used. Wild-type and MYL9 knockout samples were subjected to SDS-PAGE. ab191393 and Anti-GAPDH antibody [6C5] - Loading Control (ab8245) 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-MYL9 antibody [EPR13012(2)] (<a href='/products/primary-antibodies/myl9-antibody-epr130122-ab191393'>ab191393</a>) at 1/1000 dilution

Lane 1:

Wild-type HeLa cell lysate at 20 µg

Lane 2:

Western blot - Human MYL9 knockout HeLa cell lysate (<a href='/products/cell-lysates/human-myl9-knockout-hela-cell-lysate-ab256999'>ab256999</a>) at 20 µg

Lane 3:

Human colon cell lysate at 20 µg

Lane 4:

Daudi cell lysate at 20 µg

Secondary

Lanes 1 - 4:

Western blot - Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed (<a href='/products/secondary-antibodies/goat-rabbit-igg-h-l-irdye-800cw-preadsorbed-ab216773'>ab216773</a>) at 1/20000 dilution

Lanes 1 - 4:

Western blot - Goat anti-Mouse IgG H&L (IRDye® 680RD) preadsorbed (<a href='/products/secondary-antibodies/goat-mouse-igg-h-l-irdye-680rd-preadsorbed-ab216776'>ab216776</a>) at 1/20000 dilution

Predicted band size: 20 kDa

Observed band size: 20 kDa,37 kDa

false

• WB

Lab

Western blot - Anti-GAPDH antibody [6C5] - Loading Control (AB8245)

False colour image of Western blot : Anti-RPE65 antibody [EPR22579-44] staining at 1/1000 dilution, shown in green; Mouse anti-GAPDH antibody [6C5] (ab8245) loading control staining at 1/20000 dilution, shown in red. In Western blot, ab231782 was shown to bind specifically to RPE65. 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-RPE65 antibody [EPR22579-44] (<a href='/products/primary-antibodies/rpe65-antibody-epr22579-44-ab231782'>ab231782</a>) at 1/1000 dilution

Lane 1:

Human Eye cell lysate at 20 µg

Lane 2:

Mouse Eye cell lysate at 20 µg

Lane 3:

Rat Eye cell lysate at 20 µg

Lane 4:

Y79 cell lysate at 20 µg

Lane 5:

PC-3 cell lysate at 20 µg

Observed band size: 65 kDa

false

• WB

Lab

Western blot - Anti-GAPDH antibody [6C5] - Loading Control (AB8245)

This blot was produced using a 4-12% Bis-tris gel under the MES 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 before being incubated with ab140751 overnight at 4°C. Antibody binding was detected using the Donkey Anti-Rabbit IgG H&L (IRDye^® 680RD) preadsorbed ab216779 at a 1 : 10,000 dilution for 1hr at room temperature and then imaged using the Licor Odyssey CLx.

All lanes:

Western blot - Anti-GAPDH antibody [6C5] - Loading Control (ab8245) at 1/2000 dilution

Lane 1:

Wild-type HAP1 cell lysate (20 µg)

Lane 2:

NF-κB p60 knockout HAP1 cell lysate (20 µg)

Lane 3:

HeLa cell lysate (20 µg)

Lane 4:

A431 cell lysate (20 µg)

Secondary

All lanes:

Western blot - Donkey Anti-Rabbit IgG H&L (IRDye® 680RD) preadsorbed (<a href='/products/secondary-antibodies/donkey-rabbit-igg-h-l-irdye-680rd-preadsorbed-ab216779'>ab216779</a>) at 1/10000 dilution

Predicted band size: 36 kDa

false

• WB

Lab

Western blot - Anti-GAPDH antibody [6C5] - Loading Control (AB8245)

Lanes 1 - 4 : Merged signal (red and green). Green - ab157464 observed at 33 kDa. Red - loading control, ab8245, observed at 37 kDa.

ab157464 was shown to recognize MBD3 when MBD3 knockout samples were used, along with additional cross-reactive bands. Wild-type and MBD3 knockout samples were subjected to SDS-PAGE. ab157464 and ab8245 (loading control to GAPDH) were diluted 1/1000 and 1/10000 respectively and incubated overnight at 4°C. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye^® 800CW) preadsorbed and Goat anti-Mouse IgG H&L (IRDye^® 680RD) preadsorbed secondary antibodies at 1/10000 dilution for 1 hour at room temperature before imaging.

All lanes:

Western blot - Anti-MBD3 antibody [EPR9913] - ChIP Grade (<a href='/products/primary-antibodies/mbd3-antibody-epr9913-chip-grade-ab157464'>ab157464</a>) at 1/1000 dilution

Lanes 1 and 3:

Wild-type HAP1 cell lysate at 20 µg

Lanes 2 and 4:

MBD3 knockout HAP1 cell lysate at 20 µg

Secondary

Lanes 1 - 4:

Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed at 1/10000 dilution

Lanes 1 - 4:

Goat anti-Mouse IgG H&L (IRDye® 680RD) preadsorbed at 1/10000 dilution

Predicted band size: 33 kDa

Observed band size: 33 kDa

false

• WB

Lab

Western blot - Anti-GAPDH antibody [6C5] - Loading Control (AB8245)

Western blot : Rabbit Monoclonal [EPR28693-84] to LIF ab320821 staining at 1/1000 dilution, shown in green; Mouse anti GAPDH ab8245 loading control staining at 1/20000 dilution, shown in magenta. A band was observed at 65-80 kDa. The identity of the band detected at ~30kDa is unknown. To generate this image, samples were run on an SDS-PAGE gel then transferred onto a nitrocellulose membrane. Membranes were blocked in 3pc 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-Rabbit 800CW and Goat anti-Mouse 680RD at 1/20,000 dilution.

All lanes:

Western blot - Anti-LIF antibody [EPR28693-84] (<a href='/products/primary-antibodies/lif-antibody-epr28693-84-ab320821'>ab320821</a>) at 1/1000 dilution

Lane 1:

THP-1 at 20 µg

Lane 2:

PANC-1 at 20 µg

Lane 3:

PANC-1 Supernatant at 15 µL

Lane 4:

BT-549 at 20 µg

Lane 5:

BT-549 Supernatant at 15 µL

Secondary

All lanes:

Goat anti-Rabbit 800CW & Goat anti-Mouse 680RD at 1/20000 dilution

Predicted band size: 22 kDa

Observed band size: 65-90 kDa,37 kDa

false

• WB

Lab

Western blot - Anti-GAPDH antibody [6C5] - Loading Control (AB8245)

Western blot : Anti-CNOT6 antibody staining at 1/1000 dilution, shown in green; Mouse anti GAPDH ab8245 loading control staining at 1/20000 dilution, shown in magenta. A band was observed at 63 kDa in Wild-type MCF7 cell lysates with no signal observed at this size in CNOT6 knockout MCF7 cell line. To generate this image, samples were run on an SDS-PAGE gel then transferred onto a nitrocellulose membrane. Membranes were blocked in 3pc 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-Rabbit 800CW and Goat anti-Mouse 680RD at 1/20,000 dilution.

All lanes:

CNOT6 (E1L8F) Rabbit mAb at 1/1000 dilution

Lane 1:

Wild-type MCF7 at 20 µg

Lane 2:

Western blot - Human CNOT6 knockout MCF7 cell line (ab289386) at 20 µg

Lane 3:

HL-60 at 20 µg

Lane 4:

PC-3 at 20 µg

Secondary

All lanes:

Goat anti-Rabbit 800CW & Goat anti-Mouse 680RD at 1/20000 dilution

Predicted band size: 63 kDa

Observed band size: 63 kDa,37 kDa

false

• WB

Lab

Western blot - Anti-GAPDH antibody [6C5] - Loading Control (AB8245)

Western blot : Anti-SHC antibody [EP332Y] ab33770 staining at 1/1000 dilution, shown in green; Mouse anti GAPDH ab8245 loading control staining at 1/20000 dilution, shown in magenta. A band was observed at 50, 55, 100-140 kDa in Wild-type A549 cell lysates with no signal observed at this size in SHC1 knockout A549 cell line. To generate this image, samples were run on an SDS-PAGE gel then transferred onto a nitrocellulose membrane. Membranes were blocked in 3pc 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-Rabbit 800CW and Goat anti-Mouse 680RD at 1/20,000 dilution.

All lanes:

Western blot - Anti-SHC antibody [EP332Y] (<a href='/products/primary-antibodies/shc-antibody-ep332y-ab33770'>ab33770</a>) at 1/1000 dilution

Lane 1:

Wild-type A549 at 20 µg

Lane 2:

SHC1 knockout A549 at 20 µg

Lane 3:

HT1080 at 20 µg

Lane 4:

Raji at 20 µg

Lane 5:

HL-60 at 20 µg

Secondary

All lanes:

Goat anti-Rabbit 800CW & Goat anti-Mouse 680RD at 1/20000 dilution

Predicted band size: 100-140 kDa

Observed band size: 55 kDa,50 kDa,63 kDa,37 kDa

false

• WB

Lab

Western blot - Anti-GAPDH antibody [6C5] - Loading Control (AB8245)

Western blot : Rabbit Monoclonal[EPR3014(2)] to GGA1 ab170956 staining at 2 µg/mL, shown in green; Mouse anti GAPDH ab8245 loading control staining at 1/20000 dilution, shown in magenta. A band was observed at 74 kDa in Wild-type U-87 MG cell lysates with no signal observed at this size in GGA1 knockout U-87 MG cell line. To generate this image, samples were run on an SDS-PAGE gel then transferred onto a nitrocellulose membrane. Membranes were blocked in 3pc 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-Rabbit 800CW and Goat anti-Mouse 680RD at 1/20,000 dilution.

All lanes:

Western blot - Anti-GGA1 antibody [EPR3014(2)] (<a href='/products/primary-antibodies/gga1-antibody-epr30142-ab170956'>ab170956</a>) at 2 µg/mL

Lane 1:

Wild-type U-87 MG at 20 µg

Lane 2:

GGA1 knockout U-87 MG at 20 µg

Lane 3:

SH-SY5Y at 20 µg

Lane 4:

SK-OV-3 at 20 µg

Lane 5:

DU 145 at 20 µg

Secondary

All lanes:

Goat anti-Rabbit 800CW & Goat anti-Mouse 680RD at 1/20000 dilution

Predicted band size: 70 kDa

Observed band size: 74 kDa,36 kDa

false

• WB

Lab

Western blot - Anti-GAPDH antibody [6C5] - Loading Control (AB8245)

Lanes 1- 2 : Merged signal (red and green). Green - ab109440 observed at 120 kDa. Red - Anti-GAPDH antibody [6C5] - Loading Control (ab8245) observed at 37 kDa.

ab109440 was shown to react with NFkB p100/NFKB2 in wild-type HepG2 cells in western blot. The band observed in CRISPR/Cas9 edited cell line ab262323 (CRISPR/Cas9 edited cell lysate ab257247) lane below 97kDa may represent truncated forms and cleaved fragments. This has not been investigated further. Wild-type HepG2 and NFKB2 CRISPR/Cas9 edited HepG2 cell lysates were subjected to SDS-PAGE. Membrane was blocked for 1 hour at room temperature in 0.1% TBST with 3% non-fat dried milk. ab109440 and Anti-GAPDH antibody [6C5] - Loading Control (ab8245) were incubated overnight at 4°C at a 1 in 1000 dilution and a 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-NFkB p100/NFKB2 antibody [EPR4686] (<a href='/products/primary-antibodies/nfkb-p100-nfkb2-antibody-epr4686-ab109440'>ab109440</a>) at 1/1000 dilution

Lane 1:

Wild-type HepG2 cell lysate at 20 µg

Lane 2:

Western blot - Human NFKB2 (NFkB p100/NFKB2) knockout Hep G2 cell lysate (<a href='/products/cell-lysates/human-nfkb2-nfkb-p100-nfkb2-knockout-hep-g2-cell-lysate-ab257247'>ab257247</a>) at 20 µg

Secondary

Lanes 1 - 2:

Western blot - Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed (<a href='/products/secondary-antibodies/goat-rabbit-igg-h-l-irdye-800cw-preadsorbed-ab216773'>ab216773</a>) at 1/20000 dilution

Lanes 1 - 2:

Western blot - Goat anti-Mouse IgG H&L (IRDye® 680RD) preadsorbed (<a href='/products/secondary-antibodies/goat-mouse-igg-h-l-irdye-680rd-preadsorbed-ab216776'>ab216776</a>) at 1/20000 dilution

Predicted band size: 30 kDa,97 kDa

Observed band size: 120 kDa,30 kDa,37 kDa

false

• WB

Lab

Western blot - Anti-GAPDH antibody [6C5] - Loading Control (AB8245)

Lanes 1-3 : Merged signal (red and green). Green - ab75973 observed at 21 kDa. Red - loading control ab8245 observed at 36 kDa.

ab75973 Anti-Ferritin heavy chain antibody [EPR3004Y] was shown to specifically react with Ferritin heavy chain in wild-type HEK293T cells. Loss of signal was observed when knockout cell line ab266581 (knockout cell lysate ab256924) was used. Wild-type and Ferritin knockout samples were subjected to SDS-PAGE. ab75973 and Anti-GAPDH antibody [6C5] - Loading Control (ab8245) were incubated at room temperature for 2. 5 hours 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-Ferritin heavy chain antibody [EPR3004Y] (<a href='/products/primary-antibodies/ferritin-heavy-chain-antibody-epr3004y-ab75973'>ab75973</a>) at 1/1000 dilution

Lane 1:

Wild-type HEK293T cell lysate at 20 µg

Lane 2:

Western blot - Human FTH1 knockout HEK-293T cell lysate (<a href='/products/cell-lysates/human-fth1-knockout-hek-293t-cell-lysate-ab256924'>ab256924</a>) at 20 µg

Lane 3:

SH-SY5Y cell lysate at 20 µg

Secondary

All lanes:

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

Predicted band size: 21 kDa

Observed band size: 21 kDa,36 kDa

false

• WB

Lab

Western blot - Anti-GAPDH antibody [6C5] - Loading Control (AB8245)

Western blot : Anti-CNOT6 antibody [EPR22022] ab221151 staining at 1/1000 dilution, shown in green; Mouse anti GAPDH ab8245 loading control staining at 1/20000 dilution, shown in magenta. A band was observed at 63 kDa in Wild-type MCF7 cell lysates with no signal observed at this size in CNOT6 knockout MCF7 cell line. To generate this image, samples were run on an SDS-PAGE gel then transferred onto a nitrocellulose membrane. Membranes were blocked in 3pc 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-Rabbit 800CW and Goat anti-Mouse 680RD at 1/20,000 dilution.

All lanes:

Western blot - Anti-CNOT6 antibody [EPR22022] (<a href='/products/primary-antibodies/cnot6-antibody-epr22022-ab221151'>ab221151</a>) at 1/1000 dilution

Lane 1:

Wild-type MCF7 at 20 µg

Lane 2:

Western blot - Human CNOT6 knockout MCF7 cell line (ab289386) at 20 µg

Lane 3:

HL-60 at 20 µg

Lane 4:

PC-3 at 20 µg

Secondary

All lanes:

Goat anti-Rabbit 800CW & Goat anti-Mouse 680RD at 1/20000 dilution

Predicted band size: 63 kDa

Observed band size: 63 kDa,37 kDa

false

• WB

Lab

Western blot - Anti-GAPDH antibody [6C5] - Loading Control (AB8245)

Western blot : Anti-STK25 antibody [EPR10306] ab157188 staining at 1/1000 dilution, shown in green; Mouse anti GAPDH ab8245 loading control staining at 1/20000 dilution, shown in magenta. A band was observed at 53 kDa in Wild-type A549 cell lysates with no signal observed at this size in STK25 knockout A549 cell line. To generate this image, samples were run on an SDS-PAGE gel then transferred onto a nitrocellulose membrane. Membranes were blocked in 3pc 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-Rabbit 800CW and Goat anti-Mouse 680RD at 1/20,000 dilution.

All lanes:

Western blot - Anti-STK25 antibody [EPR10306] (<a href='/products/primary-antibodies/stk25-antibody-epr10306-ab157188'>ab157188</a>) at 1/1000 dilution

Lane 1:

Wild-type A549 at 20 µg

Lane 2:

STK25 knockout A549 at 20 µg

Lane 3:

HepG2 at 20 µg

Lane 4:

HeLa at 20 µg

Lane 5:

Daudi at 20 µg

Secondary

All lanes:

Goat anti-Rabbit 800CW & Goat anti-Mouse 680RD at 1/20000 dilution

Predicted band size: 48 kDa

Observed band size: 53 kDa,37 kDa

false

• WB

Lab

Western blot - Anti-GAPDH antibody [6C5] - Loading Control (AB8245)

Western blot : Anti-PDIA6 antibody [EPR10132(B)] ab154820 staining at 1/1000 dilution, shown in green; Mouse anti GAPDH ab8245 loading control staining at 1/20000 dilution, shown in magenta. A band was observed at 49 kDa in Wild-type A549 cell lysates with no signal observed at this size in PDIA6 knockout A549 cell line. To generate this image, samples were run on an SDS-PAGE gel then transferred onto a nitrocellulose membrane. Membranes were blocked in 3pc 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-Rabbit 800CW and Goat anti-Mouse 680RD at 1/20,000 dilution.

All lanes:

Western blot - Anti-PDIA6 antibody [EPR10132(B)] (<a href='/products/primary-antibodies/pdia6-antibody-epr10132b-ab154820'>ab154820</a>) at 1/1000 dilution

Lane 1:

Wild-type A549 at 20 µg

Lane 2:

PDIA6 knockout A549 at 20 µg

Lane 3:

THP-1 at 20 µg

Lane 4:

HT1080 at 20 µg

Lane 5:

HeLa at 20 µg

Secondary

All lanes:

Goat anti-Rabbit 800CW & Goat anti-Mouse 680RD at 1/20000 dilution

Predicted band size: 48 kDa

Observed band size: 49 kDa,37 kDa

false

• WB

Lab

Western blot - Anti-GAPDH antibody [6C5] - Loading Control (AB8245)

Western blot : Rabbit Recombinant Multiclonal[RM1080] to FAP ab314456 staining at 1/1000 dilution, shown in green; Mouse anti GAPDH ab8245 loading control staining at 1/20000 dilution, shown in magenta. A band was observed at 90-100 kDa in Wild-type U-87 MG cell lysates with no signal observed at this size in FAP knockout U-87 MG cell line. To generate this image, samples were run on an SDS-PAGE gel then transferred onto a nitrocellulose membrane. Membranes were blocked in 3pc 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-Rabbit 800CW and Goat anti-Mouse 680RD at 1/20,000 dilution.

All lanes:

Western blot - Anti-FAP antibody [RM1080] (<a href='/products/primary-antibodies/fap-antibody-rm1080-ab314456'>ab314456</a>) at 1/1000 dilution

Lane 1:

Wild-type U-87 MG at 20 µg

Lane 2:

Western blot - Human FAP knockout U-87 MG cell line (ab306819) at 20 µg

Lane 3:

WI-38 at 20 µg

Lane 4:

A549 at 20 µg

Secondary

All lanes:

Goat anti-Rabbit 800CW & Goat anti-Mouse 680RD at 1/20000 dilution

Predicted band size: 88 kDa

Observed band size: 90-100 kDa,36 kDa

false

• WB

Lab

Western blot - Anti-GAPDH antibody [6C5] - Loading Control (AB8245)

Western blot : Anti-Neuraminidase antibody [EPR15712] ab197020 staining at 1/2000 dilution, shown in green; Mouse anti GAPDH ab8245 loading control staining at 1/20000 dilution, shown in magenta. A band was observed at 40-48 kDa in Wild-type A549 UNBOILED cell lysates with no signal observed at this size in NEU1 knockout A549 UNBOILED cell line. To generate this image, samples were run on an SDS-PAGE gel then transferred onto a nitrocellulose membrane. Membranes were blocked in 5pc BSA 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-Rabbit 800CW and Goat anti-Mouse 680RD at 1/20,000 dilution.

All lanes:

Western blot - Anti-Neuraminidase antibody [EPR15712] (<a href='/products/primary-antibodies/neuraminidase-antibody-epr15712-ab197020'>ab197020</a>) at 1/2000 dilution

Lane 1:

Wild-type A549 UNBOILED at 20 µg

Lane 2:

Western blot - Human NEU1 knockout A549 cell line (<a href='/products/cell-lines/human-neu1-knockout-a549-cell-line-ab301095'>ab301095</a>) at 20 µg

Lane 3:

HeLa UNBOILED at 20 µg

Secondary

All lanes:

Goat anti-Rabbit 800CW & Goat anti-Mouse 680RD at 1/20000 dilution

Predicted band size: 45 kDa

Observed band size: 40-48 kDa,37 kDa

false

• WB

Lab

Western blot - Anti-GAPDH antibody [6C5] - Loading Control (AB8245)

Western blot : Anti-KAP1 antibody [EPR5217] ab109289 staining at 1/1000 dilution, shown in green; Mouse anti GAPDH ab8245 loading control staining at 1/20000 dilution, shown in magenta. A band was observed at 110 kDa in Wild-type HCT 116 cell lysates with no signal observed at this size in TRIM28 knockout HCT 116 cell line. To generate this image, samples were run on an SDS-PAGE gel then transferred onto a nitrocellulose membrane. Membranes were blocked in 3pc 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-Rabbit 800CW and Goat anti-Mouse 680RD at 1/20,000 dilution.

All lanes:

Western blot - Anti-KAP1 antibody [EPR5217] (<a href='/products/primary-antibodies/kap1-antibody-epr5217-ab109289'>ab109289</a>) at 1/1000 dilution

Lane 1:

Wild-type HCT 116 at 20 µg

Lane 2:

Western blot - Human TRIM28 knockout HCT116 cell line (ab289073) at 20 µg

Lane 3:

HeLa at 20 µg

Lane 4:

A-431 at 20 µg

Secondary

All lanes:

Goat anti-Rabbit 800CW & Goat anti-Mouse 680RD at 1/20000 dilution

Predicted band size: 89 kDa

Observed band size: 110 kDa,36 kDa

false

• WB

Lab

Western blot - Anti-GAPDH antibody [6C5] - Loading Control (AB8245)

Western blot : Anti-NECTIN2 antibody staining at 1/1000 dilution, shown in green; Mouse anti GAPDH ab8245 loading control staining at 1/20000 dilution, shown in magenta. A band was observed at 64-75 kDa in Wild-type A549 UNBOILED cell lysates with no signal observed at this size in NECTIN2 knockout A549 UNBOILED cell line. To generate this image, samples were run on an SDS-PAGE gel then transferred onto a nitrocellulose membrane. Membranes were blocked in 5pc 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-Rabbit 800CW and Goat anti-Mouse 680RD at 1/20,000 dilution.

All lanes:

Nectin-2/CD112 (D8D3F) XP® Rabbit mAb at 1/1000 dilution

Lane 1:

Wild-type A549 UNBOILED at 20 µg

Lane 2:

Western blot - Human NECTIN2 knockout A549 cell line (<a href='/products/cell-lines/human-nectin2-knockout-a549-cell-line-ab288906'>ab288906</a>) at 20 µg

Lane 3:

MCF7 UNBOILED at 20 µg

Lane 4:

Raji UNBOILED at 20 µg

Secondary

All lanes:

Goat anti-Rabbit 800CW & Goat anti-Mouse 680RD at 1/20000 dilution

Predicted band size: 58 kDa

Observed band size: 64-75 kDa,36 kDa

false

• WB

Unknown

Western blot - Anti-GAPDH antibody [6C5] - Loading Control (AB8245)

False colour image of Western blot : Anti-Plakophilin 3 antibody [EPR5560] staining at 1/10000 dilution, shown in green; Mouse anti-GAPDH antibody [6C5] (ab8245) loading control staining at 1/20000 dilution, shown in red. In Western blot, ab109441 was shown to bind specifically to Plakophilin 3. A band was observed at 70 and 80 kDa in wild-type HeLa cell lysates with no signal observed at this size in PKP3 CRISPR-Cas9 edited cell line ab265539 (CRISPR-Cas9 edited cell lysate ab258120). The band observed in the CRISPR-Cas9 edited lysate lane below 70 and 80 kDa is likely to represent a truncated form of Plakophilin 3. 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 PKP3 CRISPR-Cas9 edited HeLa 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-Rabbit IgG H&L (IRDye^® 800CW) preabsorbed (ab216773) and Goat anti-Mouse IgG H&L (IRDye^® 680RD) preabsorbed (ab216776) at 1/20000 dilution.

All lanes:

Western blot - Anti-Plakophilin 3 antibody [EPR5560] (<a href='/products/primary-antibodies/plakophilin-3-antibody-epr5560-ab109441'>ab109441</a>) at 1/10000 dilution

Lane 1:

Wild-type HeLa cell lysate at 20 µg

Lane 2:

PKP3 CRISPR-Cas9 edited HeLa cell lysate at 20 µg

Lane 2:

Western blot - Human PKP3 (Plakophilin 3) knockout HeLa cell lysate (<a href='/products/cell-lysates/human-pkp3-plakophilin-3-knockout-hela-cell-lysate-ab258120'>ab258120</a>) at 20 µg

Lane 3:

A431 cell lysate at 20 µg

Lane 4:

Jurkat cell lysate at 20 µg

Secondary

Lanes 1 - 4:

Western blot - Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed (<a href='/products/secondary-antibodies/goat-rabbit-igg-h-l-irdye-800cw-preadsorbed-ab216773'>ab216773</a>) at 1/20000 dilution

Lanes 1 - 4:

Western blot - Goat anti-Mouse IgG H&L (IRDye® 680RD) preadsorbed (<a href='/products/secondary-antibodies/goat-mouse-igg-h-l-irdye-680rd-preadsorbed-ab216776'>ab216776</a>) at 1/20000 dilution

Predicted band size: 87 kDa

Observed band size: 70 kDa,80 kDa,36 kDa

false