Human RAB8A knockout HeLa cell lysate

• ICC/IF

Supplier Data

Immunocytochemistry/ Immunofluorescence - Human RAB8A knockout HeLa cell lysate (AB257195)

ab241061 was shown to react with RAB8A in wild-type HeLa cells in immunocytochemistry with loss of signal observed in RAB8A knockout cell line ab264993. Wild-type and knockout cells were mixed and pelleted at a 1 : 1 ratio on coverslips. The cells were fixed with 4% paraformaldehyde (15 min) then permeabilized with 0.1% Triton X-100 (10min) and then blocked with 1x PBS, 0.01% Triton X-100, 5% BSA, 5% NGS. The cells were then incubated with ab241061 at 1/600 dilution overnight at 4°C followed by a further incubation at room temperature for 1h with a goat anti-rabbit secondary antibody to (Alexa Fluor® 555) at 0.5 ug/ml.

Acquisition of the green (wild-type), red (antibody staining) and far-red (knockout) channels was performed. Representative grayscale images of the red channel are shown. Wild-type and knockout cells are outlined with yellow and magenta dashed line, respectively. Schematic representation of the mosaic strategy used is shown on the bottom-right panel. Image was acquired with a Zeiss(LSM-880).

This data was provided by YCharOS Inc., an open science company with the mission of characterizing commercially available antibody reagents for all human proteins. Abcam and YCharOS are working together to help address the reproducibility crisis by enabling the life science community to better evaluate commercially available antibodies.

• ICC/IF

Supplier Data

Immunocytochemistry/ Immunofluorescence - Human RAB8A knockout HeLa cell lysate (AB257195)

ab237702 was shown to react with RAB8A in wild-type HeLa cells in immunocytochemistry with loss of signal observed in RAB8A knockout cell line ab264993. Wild-type and knockout cells were mixed and pelleted at a 1 : 1 ratio on coverslips. The cells were fixed with 4% paraformaldehyde (15 min) then permeabilized with 0.1% Triton X-100 (10min) and then blocked with 1x PBS, 0.01% Triton X-100, 5% BSA, 5% NGS. The cells were then incubated with ab237702 at 1/250 dilution overnight at 4°C followed by a further incubation at room temperature for 1h with a goat anti-rabbit secondary antibody to (Alexa Fluor® 555) at 0.5 ug/ml.

Acquisition of the green (wild-type), red (antibody staining) and far-red (knockout) channels was performed. Representative grayscale images of the red channel are shown. Wild-type and knockout cells are outlined with yellow and magenta dashed line, respectively. Schematic representation of the mosaic strategy used is shown on the bottom-right panel. Image was acquired with a Zeiss(LSM-880).

This data was provided by YCharOS Inc., an open science company with the mission of characterizing commercially available antibody reagents for all human proteins. Abcam and YCharOS are working together to help address the reproducibility crisis by enabling the life science community to better evaluate commercially available antibodies.

• WB

Supplier Data

Western blot - Human RAB8A knockout HeLa cell lysate (AB257195)

ab237702 was shown to react with RAB8A in wild-type HeLa cells in Western blot with loss of signal observed in RAB8A knockout cell line ab264993. Wild-type HeLa and RAB8A knockout cell lysates were subjected to SDS-PAGE. Membranes were blocked in 5% milk in TBST for 1 hr before incubation with ab237702 overnight at 4 °C at a 1/1000 dilution. Blots were incubated with secondary antibodies at 0.2 µg/mL before imaging.

This data was provided by YCharOS Inc., an open science company with the mission of characterizing commercially available antibody reagents for all human proteins. Abcam and YCharOS are working together to help address the reproducibility crisis by enabling the life science community to better evaluate commercially available antibodies.

All lanes:

Western blot - Anti-RAB8A antibody [MJF-R22] (<a href='/en-us/products/primary-antibodies/rab8a-antibody-mjf-r22-ab237702'>ab237702</a>) at 1/1000 dilution

Lane 1:

Wild-type HeLa lysate at 30 µg

Lane 2:

RAB8A knock-out HeLa lysate at 30 µg

Lane 2:

Western blot - Human RAB8A knockout HeLa cell line (<a href='/en-us/products/cell-lines/human-rab8a-knockout-hela-cell-line-ab264993'>ab264993</a>)

false

• WB

Unknown

Western blot - Human RAB8A knockout HeLa cell lysate (AB257195)

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

Lane 2 : RAB8A knockout HeLa cell lysate (20 μg)

Lane 3 : HCT116 cell lysate (20 μg)

Lane 4 : Human brain tissue lysate (20 μg)

Lanes 1-4 : Merged signal (red and green). Green - ab241061 observed at 24 kDa. Red - loading control, ab8245 observed at 37 kDa.

ab241061 Anti-RAB8A antibody [MJF-R22-79-3] was shown to specifically react with RAB8A in wild-type HeLa cells. Loss of signal was observed when knockout cell line ab264993 (knockout cell lysate ab257195) was used. Wild-type and RAB8A knockout samples were subjected to SDS-PAGE. ab ab241061 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-RAB8A antibody [MJF-R22-79-3] (<a href='/en-us/products/primary-antibodies/rab8a-antibody-mjf-r22-79-3-ab241061'>ab241061</a>) at 1/1000 dilution

Lane 1:

Wild-type HeLa cell lysate at 20 µg

Lane 2:

RAB8A knockout HeLa cell lysate at 20 µg

Lane 3:

HCT116 cell lysate at 20 µg

Lane 4:

Human brain tissue lysate at 20 µg

Predicted band size: 24 kDa

Observed band size: 24 kDa

false

• WB

Supplier Data

Western blot - Human RAB8A knockout HeLa cell lysate (AB257195)

ab241061 was shown to react with RAB8A in wild-type HeLa cells in Western blot with loss of signal observed in RAB8A knockout cell line ab264993. Wild-type HeLa and RAB8A knockout cell lysates were subjected to SDS-PAGE. Membranes were blocked in 5% milk in TBST for 1 hr before incubation with ab241061 overnight at 4 °C at a 1/1000 dilution. Blots were incubated with secondary antibodies at 0.2 µg/mL before imaging.

This data was provided by YCharOS Inc., an open science company with the mission of characterizing commercially available antibody reagents for all human proteins. Abcam and YCharOS are working together to help address the reproducibility crisis by enabling the life science community to better evaluate commercially available antibodies.

All lanes:

Western blot - Anti-RAB8A antibody [MJF-R22-79-3] (<a href='/en-us/products/primary-antibodies/rab8a-antibody-mjf-r22-79-3-ab241061'>ab241061</a>) at 1/1000 dilution

Lane 1:

Wild-type HeLa lysate at 30 µg

Lane 2:

RAB8A knock-out HeLa lysate at 30 µg

Lane 2:

Western blot - Human RAB8A knockout HeLa cell line (<a href='/en-us/products/cell-lines/human-rab8a-knockout-hela-cell-line-ab264993'>ab264993</a>)

false

• WB

Supplier Data

Western blot - Human RAB8A knockout HeLa cell lysate (AB257195)

ab188574 was shown to react with RAB8A in wild-type HeLa cells in Western blot with loss of signal observed in RAB8A knockout cell line ab264993. Wild-type HeLa and RAB8A knockout cell lysates were subjected to SDS-PAGE. Membranes were blocked in 5% milk in TBST for 1 hr before incubation with ab188574 overnight at 4 °C at a 1/1000 dilution. Blots were incubated with secondary antibodies at 0.2 µg/mL before imaging.

This data was provided by YCharOS Inc., an open science company with the mission of characterizing commercially available antibody reagents for all human proteins. Abcam and YCharOS are working together to help address the reproducibility crisis by enabling the life science community to better evaluate commercially available antibodies.

All lanes:

Western blot - Anti-RAB8A antibody [EPR14873] (<a href='/en-us/products/primary-antibodies/rab8a-antibody-epr14873-ab188574'>ab188574</a>) at 1/1000 dilution

Lane 1:

Wild-type HeLa lysate at 30 µg

Lane 2:

RAB8A knock-out HeLa lysate at 30 µg

Lane 2:

Western blot - Human RAB8A knockout HeLa cell line (<a href='/en-us/products/cell-lines/human-rab8a-knockout-hela-cell-line-ab264993'>ab264993</a>)

false

• WB

Unknown

Western blot - Human RAB8A knockout HeLa cell lysate (AB257195)

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

Lane 2 : RAB8A knockout HeLa cell lysate (20 μg)

Lane 3 : HCT116 cell lysate (20 μg)

Lane 4 : Human brain tissue lysate (20 μg)

Lanes 1-4 : Merged signal (red and green). Green - ab188574 observed at 24 kDa. Red - loading control, ab8245 observed at 37 kDa.

ab188574 Anti-RAB8A antibody [EPR14873] was shown to specifically react with RAB8A in wild-type HeLa cells. Loss of signal was observed when knockout cell line ab264993 (knockout cell lysate ab257195) was used. Wild-type and RAB8A knockout samples were subjected to SDS-PAGE. ab188574 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-RAB8A antibody [EPR14873] (<a href='/en-us/products/primary-antibodies/rab8a-antibody-epr14873-ab188574'>ab188574</a>) at 1/1000 dilution

Lane 1:

Wild-type HeLa cell lysate at 20 µg

Lane 2:

RAB8A knockout HeLa cell lysate at 20 µg

Lane 3:

HCT116 cell lysate at 20 µg

Lane 4:

Human brain tissue lysate at 20 µg

Predicted band size: 24 kDa

Observed band size: 24 kDa

false

• Sanger seq

Unknown

Sanger Sequencing - Human RAB8A knockout HeLa cell lysate (AB257195)

Homozygous : 423 bp deletion in exon 1