Anti-CIRP antibody [EPR18783]

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-CIRP antibody [EPR18783] (AB191885)

Immunohistochemical analysis of paraffin-embedded human endometrium tissue labeling CIRP using ab191885 at 1/1000 dilution. A Goat Anti-Rabbit IgG H&L (HRP) (ab97051) was used as secondary at 1/500 dilution. Counterstain : Hematoxylin.
Inset image : negative control obtained using PBS instead of ab191885, and secondary antibody only.
Note : Nuclear staining on normal human endometrium.

Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-CIRP antibody [EPR18783] (AB191885)

Immunohistochemical analysis of paraffin-embedded human breast cancer tissue labeling CIRP using ab191885 at 1/1000 dilution. A Goat Anti-Rabbit IgG H&L (HRP) (ab97051) was used as secondary at 1/500 dilution. Counterstain : Hematoxylin.
Inset image : negative control obtained using PBS instead of ab191885, and secondary antibody only.
Note : Nuclear and weak cytoplasm staining on tumor cells of breast cancer.
Mol Carcinog. 2010 49, 130–140.

Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.

• ICC/IF

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Immunocytochemistry/ Immunofluorescence - Anti-CIRP antibody [EPR18783] (AB191885)

Immunofluorescent analysis of 4% paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized HeLa (Human epithelial cells from cervix adenocarcinoma) cells labeling CIRP with ab191885 at 1/1000 dilution, followed by Goat anti-rabbit IgG (Alexa Fluor® 488) (ab150077) secondary antibody at 1/1000 dilution (green).

Confocal image showing nuclear staining on HeLa cell line.

The nuclear counter stain is DAPI (blue).

Tubulin is detected with ab7291 (anti-Tubulin mouse mAb) at 1/1000 dilution and ab150120 (AlexaFluor®594 Goat anti-Mouse secondary) at 1/1000 dilution (red).
The negative controls are as follows :
1. ab191885 at 1/1000 dilution followed by ab150120 (AlexaFluor®594 Goat anti-Mouse secondary) at 1/1000 dilution.
2. ab7291 (anti-Tubulin mouse mAb) at 1/1000 dilution followed by ab150077 (Alexa Fluor®488 Goat Anti-Rabbit IgG H&L) at 1/1000 dilution.

• ICC/IF

Supplier Data

Immunocytochemistry/ Immunofluorescence - Anti-CIRP antibody [EPR18783] (AB191885)

Immunofluorescent analysis of 4% paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized HCT 116 (Human colorectal carcinoma cell line) cells labeling CIRP with ab191885 at 1/1000 dilution, followed by Goat anti-rabbit IgG (Alexa Fluor® 488) (ab150077) secondary antibody at 1/1000 dilution (green).

Confocal image showing nuclear staining on HCT 116 cell line.

The nuclear counter stain is DAPI (blue).

Tubulin is detected with ab7291 (anti-Tubulin mouse mAb) at 1/1000 dilution and ab150120 (AlexaFluor®594 Goat anti-Mouse secondary) at 1/1000 dilution (red).
The negative controls are as follows :
1. ab191885 at 1/1000 dilution followed by ab150120 (AlexaFluor®594 Goat anti-Mouse secondary) at 1/1000 dilution.
2. ab7291 (anti-Tubulin mouse mAb) at 1/1000 dilution followed by ab150077 (Alexa Fluor®488 Goat Anti-Rabbit IgG H&L) at 1/1000 dilution.

• IP

Supplier Data

Immunoprecipitation - Anti-CIRP antibody [EPR18783] (AB191885)

Immunoprecipitation of CIRP from 1mg of HeLa (Human epithelial cells from cervix adenocarcinoma) whole cell lysate achieved using ab191885 at 1/20 dilution.
Lane 1 : Input : 10μg of HeLa whole cell lysate.
Lane 2 : HeLa whole cell lysate following IP with ab191885.
Lane 3 : negative control : IP using Rabbit monoclonal IgG (ab172730) instead of ab191885 in HeLa whole cell lysates.
Western blot was performed using ab191885 at 1/1000 dilution.
VeriBlot for IP Detection Reagent (HRP) (ab131366) at 1/10000 was used for detection.
Blocking and dilution buffer and concentration : 5% NFDM/TBST. 3 second exposure.

All lanes:

Immunoprecipitation - Anti-CIRP antibody [EPR18783] (ab191885)

Predicted band size: 18 kDa

Observed band size: 19 kDa

false

• WB

Supplier Data

Western blot - Anti-CIRP antibody [EPR18783] (AB191885)

5% NFDM/TBST : Blocking and diluting buffer.

All lanes:

Western blot - Anti-CIRP antibody [EPR18783] (ab191885) at 1/1000 dilution

Lane 1:

Human fetal heart lysate at 10 µg

Lane 2:

Human fetal kidney lysate at 10 µg

Secondary

All lanes:

Anti-Rabbit IgG (HRP), specific to the non-reduced form of IgG at 1/10000 dilution

Predicted band size: 18 kDa

Observed band size: 19 kDa

false

Exposure time: 3min

• WB

Lab

Western blot - Anti-CIRP antibody [EPR18783] (AB191885)

Lanes 1-2 : Merged signal (red and green). Green - ab191885 observed at 19 kDa. Red - loading control ab8245 observed at 37 kDa.

ab191885 Anti-CIRP antibody [EPR18783] was shown to specifically react with CIRP in wild-type HEK-293T cells. Loss of signal was observed when knockout cell line ab266187 (knockout cell lysate ab257252) was used. Wild-type and CIRP knockout samples were subjected to SDS-PAGE. ab191885 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-CIRP antibody [EPR18783] (ab191885) at 1/1000 dilution

Lane 1:

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

Lane 2:

CIRP knockout HEK-293T cell lysate at 20 µg

Lane 2:

Western blot - Human CIRBP (CIRP) knockout HEK-293T cell line (<a href='/en-us/products/cell-lines/human-cirbp-cirp-knockout-hek-293t-cell-line-ab266187'>ab266187</a>)

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 1/20000 dilution

Predicted band size: 18 kDa

Observed band size: 19 kDa

false

• WB

Supplier Data

Western blot - Anti-CIRP antibody [EPR18783] (AB191885)

5% NFDM/TBST : Blocking and diluting buffer.

The expression profile observed is consistent with what has been described in the literature (PMID : 20735994)

All lanes:

Western blot - Anti-CIRP antibody [EPR18783] (ab191885) at 1/1000 dilution

Lane 1:

HeLa (Human epithelial cells from cervix adenocarcinoma) whole cell lysate treated at 37°C for 48 hours at 20 µg

Lane 2:

HeLa whole cell lysate treated at 32°C for 48 hours at 20 µg

Lane 3:

HCT 116 (Human colorectal carcinoma cell line) whole cell lysate at 20 µg

Lane 4:

HepG2 (Human liver hepatocellular carcinoma) whole cell lysate at 20 µg

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 1/100000 dilution

Predicted band size: 18 kDa

Observed band size: 19 kDa

false

Exposure time: 3min