Anti-TMPRSS2 antibody [EPR3862]

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-TMPRSS2 antibody [EPR3862] (AB109131)

ab109131 staining TMPRSS2 in Human prostate adenocarcinoma tissue sections by Immunohistochemistry (IHC-P - paraformaldehyde-fixed, paraffin-embedded sections). Tissue was fixed and paraffin-embedded, antigen retrieval was by heat mediation in Tris/EDTA buffer pH9. Samples were incubated with primary antibody (1/1000). An undiluted HRP-conjugated mouse anti-rabbit IgG was used as the secondary antibody. Tissue counterstained with Hematoxylin. PBS was used in the negative control rather than the Primary antibody.

• IHC-P

Unknown

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-TMPRSS2 antibody [EPR3862] (AB109131)

Immunohistochemical analysis of paraffin-embedded Human prostatic adenocarcinoma using ab109131, unpurified, at a dilution of 1/500.

Perform heat mediated antigen retrieval before commencing with IHC staining protocol.

• WB

Lab

Western blot - Anti-TMPRSS2 antibody [EPR3862] (AB109131)

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

54kDa represents full-length TMPRSS2 and 25kDa is the cleaved form. The molecular weights observed are consistent with what have been described in the literature (PMID : 11245484).

All lanes:

Western blot - Anti-TMPRSS2 antibody [EPR3862] (ab109131) at 1/1000 dilution

All lanes:

LNCaP (Human prostate carcinoma epithelial cell) 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 1/20000 dilution

Predicted band size: 54 kDa

Observed band size: 25 kDa,54 kDa

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Exposure time: 7s

• WB

Lab

Western blot - Anti-TMPRSS2 antibody [EPR3862] (AB109131)

False colour image of Western blot : Anti-TMPRSS2 antibody [EPR3862] staining at 1/2000 dilution, shown in green; Mouse anti-GAPDH antibody [6C5] (ab8245) loading control staining at 1/20000 dilution, shown in red.

In Western blot, ab109131 was shown to bind specifically to TMPRSS2. A band was observed at 55, 25 kDa in wild-type LNCaP and at 25 kDa in Caco-2 cell lysates with no signal observed at this size in TMPRSS2 knockout LNCaP cell line ab273745 (knockout LNCaP cell lysate ab275499) and TMPRSS2 knockout Caco-2 cell line ab273737 (knockout Caco-2 cell lysate ab277340).

To generate this image, wild-type and TMPRSS2 knockout cell lysates were analysed. First, samples were run on an SDS-PAGE gel then transferred onto a nitrocellulose membrane. Membranes were blocked in 5 % 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-TMPRSS2 antibody [EPR3862] (ab109131) at 1/2000 dilution

Lane 1:

Wild-type LNCaP cell lysate at 20 µg

Lane 2:

TMPRSS2 knockout LNCaP cell lysate at 20 µg

Lane 2:

Western blot - Human TMPRSS2 knockout LNCaP cell line (<a href='/en-us/products/cell-lines/human-tmprss2-knockout-lncap-cell-line-ab273745'>ab273745</a>)

Lane 3:

Wild-type Caco-2 cell lysate at 20 µg

Lane 4:

TMPRSS2 knockout Caco-2 cell lysate at 20 µg

Lane 5:

PC-3 cell lysate at 20 µg

Lane 6:

DU 145 cell lysate at 20 µg

Lane 7:

Human Prostate cell lysate at 20 µg

Lane 8:

Human Colon cell lysate at 20 µg

Lane 9:

Human Lung cell lysate at 20 µg

Predicted band size: 54 kDa

Observed band size: 25 kDa,55 kDa

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