Anti-CD44 antibody [EPR1013Y]

• IHC-P

Unknown

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-CD44 antibody [EPR1013Y] (AB51037)

ab51037 (1 : 100) showing positive staining in human Glioma tissue.

Perform heat mediated antigen retrieval with citrate buffer pH 6 before commencing with IHC staining protocol.

• IHC-P

Unknown

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-CD44 antibody [EPR1013Y] (AB51037)

Immunohistochemical analysis of paraffin-embedded human cervical cancer tissue labeling CD44 with ab51037 at 1/100 dilution followed by goat anti-rabbit IgG H&L (HRP) (ab97051, 1/500). The sample was counter stained with hematoxylin.

Perform heat mediated antigen retrieval with citrate buffer pH 6 before commencing with IHC staining protocol.

• IHC-P

Unknown

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-CD44 antibody [EPR1013Y] (AB51037)

Immunohistochemical analysis of paraffin-embedded human pancreatic cancer tissue labeling CD44 with ab51037 at 1/100 dilution followed by goat anti-rabbit IgG H&L (HRP) (ab97051, 1/500). The sample was counter stained with hematoxylin.

Perform heat mediated antigen retrieval with citrate buffer pH 6 before commencing with IHC staining protocol.

• IHC-P

Unknown

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-CD44 antibody [EPR1013Y] (AB51037)

Immunohistochemical analysis of paraffin-embedded human skin tissue labeling CD44 with ab51037 at 1/100 dilution followed by goat anti-rabbit IgG H&L (HRP) (ab97051, 1/500). The sample was counter stained with hematoxylin.

Perform heat mediated antigen retrieval with citrate buffer pH 6 before commencing with IHC staining protocol.

• IHC-P

Unknown

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-CD44 antibody [EPR1013Y] (AB51037)

ab51037 (1 : 100) showing positive staining in human Breast carcinoma tissue.

Perform heat mediated antigen retrieval with citrate buffer pH 6 before commencing with IHC staining protocol.

• IHC-P

Unknown

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-CD44 antibody [EPR1013Y] (AB51037)

Immunohistochemical analysis of paraffin-embedded human tonsil tissue labeling CD44 with ab51037 at 1/100 dilution followed by goat anti-rabbit IgG H&L (HRP) (ab97051, 1/500). The sample was counter stained with hematoxylin.

Perform heat mediated antigen retrieval with citrate buffer pH 6 before commencing with IHC staining protocol.

• IHC-P

Unknown

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-CD44 antibody [EPR1013Y] (AB51037)

ab51037 (1 : 100) showing positive staining in human Thyroid gland carcinoma tissue.

Perform heat mediated antigen retrieval with citrate buffer pH 6 before commencing with IHC staining protocol.

• WB

Lab

Western blot - Anti-CD44 antibody [EPR1013Y] (AB51037)

Lanes 1 - 4 : Merged signal (red and green). Green - ab51037 observed at 80 kDa. Red - loading control, ab8245 (Mouse anti-GAPDH antibody [6C5]) observed at 37kDa.

ab51037 was shown to react with CD44 in wild-type HeLa cells in western blot. Loss of signal was observed when CD44 knockout sample was used. Wild-type HeLa and CD44 knockout cell lysates were subjected to SDS-PAGE. Membranes were blocked in 3% milk in TBS-T (0.1% Tween^®) before incubation with ab51037 and ab8245 (Mouse anti-GAPDH antibody [6C5]) overnight at 4°C at a 1 in 5000 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-CD44 antibody [EPR1013Y] (ab51037) at 1/5000 dilution

Lane 1:

Wild-type HeLa (Human epithelial cell line from cervix adenocarcinoma) whole cell lysate at 20 µg

Lane 2:

CD44 knockout HeLa (Human epithelial cell line from cervix adenocarcinoma) whole cell lysate at 20 µg

Lane 2:

Western blot - Human CD44 knockout HeLa cell line (<a href='/en-us/products/cell-lines/human-cd44-knockout-hela-cell-line-ab262515'>ab262515</a>)

Lane 3:

A549 (Human lung carcinoma cell line) whole cell lysate at 20 µg

Lane 4:

LNCaP (Human prostate cancer cell line) whole cell lysate at 20 µg

Predicted band size: 81 kDa

Observed band size: 80 kDa

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• WB

Lab

Western blot - Anti-CD44 antibody [EPR1013Y] (AB51037)

False colour image of Western blot : Anti-CD44 antibody [EPR1013Y] staining at 1/5000 dilution, shown in green; Mouse anti-Alpha Tubulin [DM1A] (ab7291) loading control staining at 1/20000 dilution, shown in red. In Western blot, ab51037 was shown to bind specifically to CD44. A band was observed at 75-80 kDa in wild-type HeLa cell lysates with no signal observed at this size in CD44 knockout cell line ab262515 (knockout cell lysate ab263938). To generate this image, wild-type and CD44 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-CD44 antibody [EPR1013Y] (ab51037) at 1/5000 dilution

Lane 1:

Wild-type HeLa cell lysate at 20 µg

Lane 2:

CD44 knockout HeLa cell lysate at 20 µg

Lane 2:

Western blot - Human CD44 knockout HeLa cell line (<a href='/en-us/products/cell-lines/human-cd44-knockout-hela-cell-line-ab262515'>ab262515</a>)

Lane 3:

A549 cell lysate at 20 µg

Lane 4:

LNCaP cell lysate at 20 µg

Predicted band size: 81 kDa

Observed band size: 75-80 kDa

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• WB

Lab

Western blot - Anti-CD44 antibody [EPR1013Y] (AB51037)

Blocking and diluting buffer 5% NFDM/TBST

The expression of CD44 in MCF-7 is low (PMID : 25635866; PMID : 26005723). Jurkat does not express CD44 (PMID : 24127558).

All lanes:

Western blot - Anti-CD44 antibody [EPR1013Y] (ab51037) at 1/1000 dilution

Lane 1:

MCF-7 whole cell lysate at 10 µg

Lane 2:

Jurkat whole cell lysate at 10 µg

Lane 3:

MDA-MB-231 whole cell lysate at 10 µg

Lane 4:

HeLa whole cell lysate at 10 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 1/20000 dilution

Predicted band size: 81 kDa

Observed band size: 81 kDa

false

Exposure time: 3min

• WB

Unknown

Western blot - Anti-CD44 antibody [EPR1013Y] (AB51037)

All lanes:

Western blot - Anti-CD44 antibody [EPR1013Y] (ab51037) at 1/5000 dilution

All lanes:

TF-1 cell lysate at 10 µg

Secondary

All lanes:

Goat anti-Rabbit HRP labeled. at 1/2000 dilution

Predicted band size: 81 kDa

Observed band size: 82 kDa

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• OI-RD Scanning

Unknown

OI-RD Scanning - Anti-CD44 antibody [EPR1013Y] (AB51037)

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.