重组Anti-Interferon alpha/beta receptor 1抗体[EPR6244]

• IHC-P

Lab

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Interferon alpha/beta receptor 1 antibody [EPR6244] (AB124764)

The immunostaining was performed on a Leica Biosystems BOND® RX instrument. Paraffin embedded human endometrium tissue slides were dewaxed, followed by heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0, Epitope Retrieval ER2 Solution) for 20 mins. Endogenous peroxidase activity was blocked with 3% H2O2 for 10 mins. The sections were then incubated with ab124764 (1/500) at room temperature for 30 mins, followed by a ready to use Rabbit specific IHC polymer detection kit HRP/DAB (ab209101). The sections were counterstained with Hematoxylin. The images were taken with a Leica Aperio AT2.

ab124767 was stored at a range of temperatures (+4°C, +22°C, +37°C) for 1 week before being tested in IHC-P. The image shows the staining intensity remains relatively constant across all storage temperatures, demonstrating that antibody activity is not affected.

• WB

Lab

Western blot - Anti-Interferon alpha/beta receptor 1 antibody [EPR6244] (AB124764)

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

All lanes:

Western blot - Anti-Interferon alpha/beta receptor 1 antibody [EPR6244] (ab124764) at 1/1000 dilution

Lane 1:

Untreated K-562 (Human chronic myelogenous leukemia lymphoblast) whole cell lysate. at 15 µg

Lane 2:

K-562 (Human chronic myelogenous leukemia lymphoblast) treated with PNGase F whole cell lysate. at 15 µg

Lane 3:

Untreated HeLa (Human cervix adenocarcinoma epithelial cell) whole cell lysate. at 15 µg

Lane 4:

HeLa (Human cervix adenocarcinoma epithelial cell) treated with PNGase F whole cell lysate 15 μg. at 15 µg

Secondary

All lanes:

Western blot - Goat Anti-Rabbit IgG H&L (HRP) (<a href='/products/secondary-antibodies/goat-rabbit-igg-h-l-hrp-ab97051'>ab97051</a>) at 1/20000 dilution

Predicted band size: 64 kDa

Observed band size: 64 kDa

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Exposure time: 180s

• WB

Unknown

Western blot - Anti-Interferon alpha/beta receptor 1 antibody [EPR6244] (AB124764)

All lanes:

Western blot - Anti-Interferon alpha/beta receptor 1 antibody [EPR6244] (ab124764) at 1/1000 dilution

Lane 1:

HeLa cell lysate at 10 µg

Lane 2:

HeLa cell lysate treated with IFN-alpha at 10 µg

Lane 3:

K562 cell lysate at 10 µg

Lane 4:

U937 cell lysate at 10 µg

Secondary

All lanes:

Goat anti-Rabbit HRP at 1/200 dilution

Predicted band size: 64 kDa

false

• WB

Lab

Western blot - Anti-Interferon alpha/beta receptor 1 antibody [EPR6244] (AB124764)

Blocking buffer : 5% NFDM/TBST
Dilution buffer : 5% NFDM/TBST

All lanes:

Western blot - Anti-Interferon alpha/beta receptor 1 antibody [EPR6244] (ab124764) at 1/1000 dilution

Lane 1:

HEK-293 whole cell lysate at 10 µg

Lane 2:

HeLa whole cell lysate at 10 µg

Lane 3:

LNCaP whole cell lysate at 10 µg

Lane 4:

SH-SY5Y whole cell lysate at 10 µg

Secondary

All lanes:

Western blot - Goat Anti-Rabbit IgG H&L (HRP) (<a href='/products/secondary-antibodies/goat-rabbit-igg-h-l-hrp-ab97051'>ab97051</a>) at 1/20000 dilution

Predicted band size: 64 kDa

Observed band size: 90-130 kDa

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• WB

Lab

Western blot - Anti-Interferon alpha/beta receptor 1 antibody [EPR6244] (AB124764)

Western blot : Anti-IFNAR1 antibody [EPR6244] (ab124764) staining at 1/1000 dilution, shown in green; Mouse anti-CANX [CANX/1543] (ab238078) loading control staining at 1/20000 dilution, shown in red. In Western blot, ab124764 was shown to bind specifically to IFNAR1. A band was observed at 116 kDa in wild-type HeLa cell lysates with no signal observed at this size in IFNAR1 knockout cell line. To generate this image, wild-type and IFNAR1 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 800CW and Goat anti-Mouse IgG H&L 680RD at 1/20000 dilution.

All lanes:

Western blot - Anti-Interferon alpha/beta receptor 1 antibody [EPR6244] (ab124764) at 1/1000 dilution

Lane 1:

Wild-type HeLa cell lysate at 20 µg

Lane 2:

Western blot - Human IFNAR1 (Interferon alpha/beta receptor 1) knockout HeLa cell line (<a href='/products/cell-lines/human-ifnar1-interferon-alpha-beta-receptor-1-knockout-hela-cell-line-ab261782'>ab261782</a>)

Lane 2:

IFNAR1 knockout HeLa cell lysate at 20 µg

Secondary

All lanes:

Goat anti-Rabbit IgG H&L 800CW and Goat anti-Mouse IgG H&L 680RD at 1/20000 dilution

Predicted band size: 64 kDa

Observed band size: 116 kDa

false

• WB

Lab

Western blot - Anti-Interferon alpha/beta receptor 1 antibody [EPR6244] (AB124764)

Blocking buffer : 5% NFDM/TBST
Dilution buffer : 5% NFDM/TBST

In mouse and rat tissue lysates this product detects a band in the region of 100 kDa, however we believe this band is non-specific and is not interferon receptor alpha as the immunogen for this antibody shares only 55% homology with the mouse and rat protein. In addition this band also migrates at a lower molecular weight than that detected in human samples.

All lanes:

Western blot - Anti-Interferon alpha/beta receptor 1 antibody [EPR6244] (ab124764) at 1/1000 dilution

Lane 1:

Mouse brain tissue lysate at 10 µg

Lane 2:

Rat brain tissue lysate at 10 µg

Predicted band size: 64 kDa

Observed band size: 90 kDa

false

• WB

Lab

Western blot - Anti-Interferon alpha/beta receptor 1 antibody [EPR6244] (AB124764)

Blocking buffer : 5% NFDM/TBST.

In mouse and rat tissue lysates this product detects a band in the region of 100 kDa, however we believe this band is non-specific and is not interferon receptor alpha as the immunogen for this antibody shares only 55% homology with the mouse and rat protein. In addition this band also migrates at a lower molecular weight than that detected in human samples.

All lanes:

Western blot - Anti-Interferon alpha/beta receptor 1 antibody [EPR6244] (ab124764) at 1/1000 dilution

Lane 1:

Mouse brain whole tissue lysate at 10 µg

Lane 2:

Mouse heart whole tissue lysate at 10 µg

Lane 3:

Mouse kidney whole tissue lysate at 10 µg

Lane 4:

Mouse spleen whole tissue lysate at 10 µg

Lane 5:

Rat brain whole tissue lysate at 10 µg

Lane 6:

Rat heart whole cell lysate at 10 µg

Lane 7:

Rat kidney whole cell lysate at 10 µg

Lane 8:

Rat spleen whole tissue lysate at 10 µg

Secondary

All lanes:

Goat anti-rabbit IgG (H+L), peroxidase conjugated at 1/2000 dilution

Predicted band size: 64 kDa

false

Exposure time: 3min

• OI-RD Scanning

Unknown

OI-RD Scanning - Anti-Interferon alpha/beta receptor 1 antibody [EPR6244] (AB124764)

We have systematically measured KD (the equilibrium dissociation constant between the antibody and its antigen), of more than 840 recombinant antibodies to assess not only their individual KD values but also to see the average affinity of antibody. Based on the comparison with published literature values for mouse monoclonal antibodies, Recombinant antibodies appear to be on average 1-2 order of magnitude higher affinity.