Anti-MSH6 antibody [EPR3945]

• IHC-P

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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-MSH6 antibody [EPR3945] (AB92471)

Immunohistochemical staining of paraffin embedded human colon with purified ab92471 at a dilution of 1/500. A pre-diluted HRP polymer for rabbit/mouse IgG was used as the secondary antibody and the sample was counter-stained with hematoxylin. Antigen retrieval was perfomed using Tris-EDTA buffer, pH 9.0. PBS was used instead of the primary antibody as the negative control, and is shown in the inset.

• IHC

Lab

Immunohistochemistry - Anti-MSH6 antibody [EPR3945] (AB92471)

Immunohistochemical analysis of formalin fixed paraffin embedded human small intestine labelling MSH6 with ab92471 at a concentration of 0.1µg/ml.

The immunostaining was performed on a Ventana DISCOVERY ULTRA (Roche Tissue Diagnostics) instrument with an OptiView DAB IHC Detection Kit. Heat mediated antigen retrieval was conducted for 32min with DISCOVERY cell conditioning solution (CC1) 100°C, pH8.5. ab92471 anti-MSH6 was incubated at 37°C for 16min.

Sections were counterstained is with Hematoxylin II. Image inset shows absence of staining in secondary antibody only control.

Customers are encouraged to optimise antigen retrieval conditions, antibody concentration, incubation times and temperature for best results in their own IHC assay workflow (automated and manual)

• IHC-P

Lab

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-MSH6 antibody [EPR3945] (AB92471)

Tissue Microarrays stained for "Anti-MSH6 antibody [EPR3945]" using "ab92471"in immunohistochemical analysis. This table provides a detailed overview of positive (tick mark) and negative (cross mark) staining per sample type tested. The sections were pre-treated using Heat mediated antigen retrieval using Bond™ Epitope Retrieval Solution 2 (pH 9.0) for 20 minutes. The sections were incubated with ab92471 for 30 mins at room temperature followed by a ready to use Rabbit specific IHC polymer detection kit HRP/DAB (ab209101). The immunostaining was performed on a Leica Biosystems BOND® RX instrument.

• IHC

Lab

Immunohistochemistry - Anti-MSH6 antibody [EPR3945] (AB92471)

Immunohistochemical analysis of formalin fixed paraffin embedded human small intestine labelling MSH6 with ab92471 at a concentration of 0.1µg/ml. The immunostaining was performed on a Leica Biosystems BOND® RX instrument with a Bond™ Polymer Refine Detection kit. Heat mediated antigen retrieval was performed with Tris-EDTA buffer (pH 9.0, Epitope Retrieval Solution 2) for 20 mins.

ab92471 anti-MSH6 antibody [EPR3945] was incubated for 15mins at room temperature. Sections were counterstained with Hematoxylin. Image inset shows absence of staining in secondary antibody only control.

Customers are encouraged to optimise antigen retrieval conditions, antibody concentration, incubation times and temperature for best results in their own IHC assay workflow (automated and manual)

• IHC-P

Lab

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-MSH6 antibody [EPR3945] (AB92471)

Tissue Microarrays stained for "Anti-MSH6 antibody [EPR3945]" using "ab92471"in immunohistochemical analysis. This table provides a detailed overview of positive (tick mark) and negative (cross mark) staining per sample type tested. The sections were pre-treated using Heat mediated antigen retrieval using Bond™ Epitope Retrieval Solution 2 (pH 9.0) for 20 minutes. The sections were incubated with ab92471 for 30 mins at room temperature followed by a ready to use Rabbit specific IHC polymer detection kit HRP/DAB (ab209101). The immunostaining was performed on a Leica Biosystems BOND® RX instrument.

• IHC-P

Unknown

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-MSH6 antibody [EPR3945] (AB92471)

Immunohistochemical staining of paraffin embedded rat liver with purified ab92471 at a dilution of 1/500. A pre-diluted HRP polymer for rabbit/mouse IgG was used as the secondary antibody and the sample was counter-stained with hematoxylin. Antigen retrieval was perfomed using Tris-EDTA buffer, pH 9.0. PBS was used instead of the primary antibody as the negative control, and is shown in the inset.

• WB

Lab

Western blot - Anti-MSH6 antibody [EPR3945] (AB92471)

Blocking buffer : 5% NFDM/TBST

Dilution buffer : 5% NFDM/TBST

All lanes:

Western blot - Anti-MSH6 antibody [EPR3945] (ab92471) at 1/2000 dilution

All lanes:

SW480 cell lysate at 10 µg

Secondary

All lanes:

HRP goat anti-rabbit (H+L) at 1/1000 dilution

Predicted band size: 153 kDa

Observed band size: 160 kDa

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• WB

Lab

Western blot - Anti-MSH6 antibody [EPR3945] (AB92471)

Blocking buffer : 5% NFDM/TBST

Dilution buffer : 5% NFDM/TBST

All lanes:

Western blot - Anti-MSH6 antibody [EPR3945] (ab92471) at 1/1000 dilution

All lanes:

Rat brain at 10 µg

Secondary

All lanes:

HRP goat anti-rabbit (H+L) at 1/1000 dilution

Predicted band size: 153 kDa

Observed band size: 160 kDa

false

• ICC/IF

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Immunocytochemistry/ Immunofluorescence - Anti-MSH6 antibody [EPR3945] (AB92471)

ab92471 staining MSH6 in HeLa cells. The cells were fixed with 4% formaldehyde (10min), permeabilized with 0.1% Triton X-100 for 5 minutes and then blocked with 1% BSA/10% normal goat serum/0.3M glycine in 0.1% PBS-Tween for 1h. The cells were then incubated with ab92471 at 1μg/ml and ab195889 at 1/250 dilution (shown in pseudocolour red) overnight at +4°C, followed by a further incubation at room temperature for 1h with a goat secondary antibody to Rabbit IgG (Alexa Fluor® 488) (ab150081) at 2 μg/ml (shown in green). Nuclear DNA was labelled in blue with DAPI.

Image was taken with a confocal microscope (Leica-Microsystems, TCS SP8).

• ICC/IF

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Immunocytochemistry/ Immunofluorescence - Anti-MSH6 antibody [EPR3945] (AB92471)

ab92471 staining MSH6 in wild-type HAP1 cells (top panel) and MSH6 knockout HAP1 cells (bottom panel). The cells were fixed with 4% formaldehyde (10min), permeabilized with 0.1% Triton X-100 for 5 minutes and then blocked with 1% BSA/10% normal goat serum/0.3M glycine in 0.1% PBS-Tween for 1h. The cells were then incubated with ab92471 at 1μg/ml and ab195889 at 1/250 dilution (shown in pseudocolour red) overnight at +4°C, followed by a further incubation at room temperature for 1h with a goat secondary antibody to Rabbit IgG (Alexa Fluor® 488) (ab150081) at 2 μg/ml (shown in green). Nuclear DNA was labelled in blue with DAPI.

Image was taken with a confocal microscope (Leica-Microsystems, TCS SP8).

• ICC/IF

AbReview29265****

Immunocytochemistry/ Immunofluorescence - Anti-MSH6 antibody [EPR3945] (AB92471)

Unpurified ab92471 (1/500) staining MSH6 in asynchronous HeLa cells (green). Cells were fixed in paraformaldehyde, permeabilised with 0.5% Triton X100/PBS and counterstained with DAPI in order to highlight the nucleus (red). For further experimental details please see abreview.

Image courtesy of an abreview submitted by Dr. Kirk McManus, Univ. of Manitoba/Cancer Care MICB, Canada

• IHC-P

Unknown

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-MSH6 antibody [EPR3945] (AB92471)

Unpurified ab92471, at a 1/100 dilution, detecting MSH6 in paraffin embedded Human colonic adenocarcinoma tissue by immunohistochemistry. Detection used HRP conjugated anti rabbit antibody.

Heat mediated antigen retrieval was performed before commencing with IHC staining protocol.

• WB

Lab

Western blot - Anti-MSH6 antibody [EPR3945] (AB92471)

Different batches of ab92471 were tested on Rat brain lysate at 0.2 µg/ml. 15 µg of lysate was loaded in each lane. Bands observed at 160 kDa.

All lanes:

Western blot - Anti-MSH6 antibody [EPR3945] (ab92471)

Predicted band size: 153 kDa

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• WB

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Western blot - Anti-MSH6 antibody [EPR3945] (AB92471)

Secondary antibody - goat anti-rabbit HRP (ab6721)

All lanes:

Western blot - Anti-MSH6 antibody [EPR3945] (ab92471) at 1/1000 dilution

Lane 1:

A431 cell lysate at 10 µg

Lane 2:

HeLa cell lysate at 10 µg

Lane 3:

SW480 cell lysate at 10 µg

Secondary

All lanes:

HRP labelled goat anti-rabbit antibody at 1/2000 dilution

Predicted band size: 153 kDa

false

• WB

Lab

Western blot - Anti-MSH6 antibody [EPR3945] (AB92471)

Blocking buffer : 5% NFDM/TBST

Dilution buffer : 5% NFDM/TBST

All lanes:

purified at 1/6000 dilution

All lanes:

SW480 cell lysate at 10 µg

Secondary

All lanes:

HRP goat anti-rabbit (H+L) at 1/1000 dilution

Predicted band size: 153 kDa

Observed band size: 160 kDa

false

• WB

Lab

Western blot - Anti-MSH6 antibody [EPR3945] (AB92471)

Lanes 1- 2 : Merged signal (red and green). Green - ab92471 observed at 160 kDa. Red - Anti-Vinculin antibody [VIN-54] observed at 124 kDa.

ab92471 was shown to react with MSH6 in wild-type HeLa cells in western blot. Loss of signal was observed when knockout cell line ab255410 (knockout cell lysate ab263763) was used. Wild-type HeLa and MSH6 knockout HeLa cell lysates were subjected to SDS-PAGE. Membrane was blocked for 1 hour at room temperature in 0.1% TBST with 3% non-fat dried milk. ab92471 and Anti-Vinculin antibody [VIN-54] overnight at 4°C at a 1 in 1000 dilution and a 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-MSH6 antibody [EPR3945] (ab92471) at 1/1000 dilution

Lane 1:

Wild-type HeLa cell lysate at 20 µg

Lane 2:

MSH6 knockout HeLa cell lysate at 20 µg

Lane 2:

Western blot - Human MSH6 knockout HeLa cell line (<a href='/en-us/products/cell-lines/human-msh6-knockout-hela-cell-line-ab255410'>ab255410</a>)

Predicted band size: 153 kDa

Observed band size: 160 kDa

false

• WB

Lab

Western blot - Anti-MSH6 antibody [EPR3945] (AB92471)

Lanes 1 - 4 : Merged signal (red and green). Green - ab92471 observed at 160 kDa. Red - loading control, ab18058, observed at 124 kDa.

ab92471 was shown to specifically react with MSH6 in wild-type HAP1 cells. No band was observed when MSH6 knockout samples were used. Wild-type and MSH6 knockout samples were subjected to SDS-PAGE. ab92471 and ab18058 (loading control to Vinculin) were diluted at 1/1000 and 1/10 000 respectively and incubated overnight at 4°C. 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 1/10 000 dilution for 1 h at room temperature before imaging.

All lanes:

Western blot - Anti-MSH6 antibody [EPR3945] (ab92471) at 1/1000 dilution

Lane 1:

Wild-type HAP1 cell lysate at 20 µg

Lane 2:

MSH6 knockout HAP1 cell lysate at 20 µg

Lane 3:

HeLa cell lysate at 20 µg

Lane 4:

A431 cell lysate at 20 µg

Predicted band size: 153 kDa

false

• WB

AbReview56726****

Western blot - Anti-MSH6 antibody [EPR3945] (AB92471)

All lanes:

Western blot - Anti-MSH6 antibody [EPR3945] (ab92471) at 1/1000 dilution

Lane 1:

HAP1 WT cell lysate at 50 µg

Lane 2:

MSH6 KO HAP1 cell lysate at 50 µg

Secondary

All lanes:

donkey anti-rabbit IgG-HRP at 1/3000 dilution

Predicted band size: 153 kDa

true

Exposure time: 5min

This image is courtesy of an Abreview by Serena Bologna.

• OI-RD Scanning

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OI-RD Scanning - Anti-MSH6 antibody [EPR3945] (AB92471)

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.