Anti-IL-8 antibody [EPR22994-255] - BSA and Azide free

• WB

Lab

Western blot - Anti-IL-8 antibody [EPR22994-255] - BSA and Azide free (AB275255)

This data was developed using the same antibody clone in a different buffer formulation (ab235584).

Lanes 1 - 6 : Merged signal (red and green). Green - ab235584 observed at 10 kDa. Red - loading control ab8245 (Mouse anti-GAPDH antibody [6C5]) observed at 37 kDa.

ab235584 was shown to react with IL-8 in wild-type PC-3 cells in Western blot with loss of signal observed in CXCL8 knockout sample. Wild-type PC-3 and CXCL8 knockout cell lysates were subjected to SDS-PAGE. Membranes were blocked in fluorescent western blot (TBS-based) blocking solution before incubation with ab235584 and ab8245 (Mouse anti-GAPDH antibody [6C5]) overnight at 4 °C at a 1 in 1000 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 h at room temperature before imaging.

All lanes:

Western blot - Anti-IL-8 antibody [EPR22994-255] (<a href='/en-us/products/primary-antibodies/il-8-antibody-epr22994-255-ab235584'>ab235584</a>) at 1/1000 dilution

Lane 1:

Wild-type PC-3 Brefeldin A (<a href='/en-us/products/biochemicals/brefeldin-a-inhibitor-of-adp-ribosylation-factor-ab120299'>ab120299</a>)-treated (5 &mu;g/ml, 5 h) cell lysate at 30 µg

Lane 2:

Wild-type PC-3 LPS-treated (2 &mu;g/ml, 6 h) with Brefeldin A (<a href='/en-us/products/biochemicals/brefeldin-a-inhibitor-of-adp-ribosylation-factor-ab120299'>ab120299</a>) (5 &mu;g/ml, 5 h) cell lysate at 30 µg

Lane 2:

Western blot - Human CXCL8 knockout PC-3 cell line (<a href='/en-us/products/cell-lines/human-cxcl8-knockout-pc-3-cell-line-ab273743'>ab273743</a>)

Lane 3:

CXCL8 knockout PC-3 Brefeldin A (<a href='/en-us/products/biochemicals/brefeldin-a-inhibitor-of-adp-ribosylation-factor-ab120299'>ab120299</a>)-treated (5 &mu;g/ml, 5 h) cell lysate at 30 µg

Lane 4:

CXCL8 knockout PC-3 LPS-treated (2 &mu;g/ml, 6 h) with Brefeldin A (<a href='/en-us/products/biochemicals/brefeldin-a-inhibitor-of-adp-ribosylation-factor-ab120299'>ab120299</a>) (5 &mu;g/ml, 5 h) cell lysate at 30 µg

Lane 5:

A431 cell lysate at 30 µg

Lane 6:

HCT116 cell lysate at 30 µg

Predicted band size: 11 kDa

Observed band size: 10 kDa

false

• WB

Lab

Western blot - Anti-IL-8 antibody [EPR22994-255] - BSA and Azide free (AB275255)

This data was developed using ab235584, the same antibody clone in a different buffer formulation.

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

This blot was developed using a higher sensitivity ECL substrate.

Exposure time : 3 minutes

All lanes:

Western blot - Anti-IL-8 antibody [EPR22994-255] (<a href='/en-us/products/primary-antibodies/il-8-antibody-epr22994-255-ab235584'>ab235584</a>) at 1/1000 dilution

Lane 1:

U-87 MG (human glioblastoma-astrocytoma epithelial cell), whole cell lysate at 40 µg

Lane 2:

U-87 MG treated with 1μM Thapsigargin for 24 hours, whole cell lysate at 40 µg

Secondary

All lanes:

Western blot - VeriBlot for IP Detection Reagent (HRP) (<a href='/en-us/products/reagents/veriblot-for-ip-detection-reagent-hrp-ab131366'>ab131366</a>) at 1/1000 dilution

Predicted band size: 11 kDa

Observed band size: 11 kDa

false

• WB

Lab

Western blot - Anti-IL-8 antibody [EPR22994-255] - BSA and Azide free (AB275255)

This data was developed using ab235584, the same antibody clone in a different buffer formulation.

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

Exposure time : 3 minutes

All lanes:

Western blot - Anti-IL-8 antibody [EPR22994-255] (<a href='/en-us/products/primary-antibodies/il-8-antibody-epr22994-255-ab235584'>ab235584</a>) at 1/1000 dilution

All lanes:

His-tagged human recombiant IL-8 protein (aa21-99) 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 1/20000 dilution

Predicted band size: 11 kDa

Observed band size: 11 kDa

false