Anti-Alkaline Phosphatase antibody [EPR27506-72] (BSA and Azide free)

• IHC

Supplier Data

Immunohistochemistry - Anti-Alkaline Phosphatase antibody [EPR27506-72] (BSA and Azide free) (AB305306)

This data was developed using ab305305, the same antibody clone in a different buffer formulation. Immunohistochemical analysis of paraffin-embedded human adrenal gland tissue labeling Alkaline Phosphatase with ab305305 at 1/2000 (0.261 µg/ml) followed by ready to use Leica DS9800 (Bond® Polymer Refine Detection) kit. Positive staining on human adrenal gland (PMID : 31563571). The section was incubated with ab305305 for 30 mins at room temperature. The immunostaining was performed on a Leica Biosystems BOND® RX instrument. Counterstained with Hematoxylin. Secondary antibody only control : Secondary antibody is ready to use Leica DS9800 (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.

• IHC

Supplier Data

Immunohistochemistry - Anti-Alkaline Phosphatase antibody [EPR27506-72] (BSA and Azide free) (AB305306)

This data was developed using ab305305, the same antibody clone in a different buffer formulation. Immunohistochemical analysis of paraffin-embedded human serous ovarian tissue labeling Alkaline Phosphatase with ab305305 at 1/2000 (0.261 µg/ml) followed by ready to use Leica DS9800 (Bond® Polymer Refine Detection) kit. Positive staining on human serous ovarian carcinoma (PMID : 30979497). The section was incubated with ab305305 for 30 mins at room temperature. The immunostaining was performed on a Leica Biosystems BOND® RX instrument. Counterstained with Hematoxylin. Secondary antibody only control : Secondary antibody is ready to use Leica DS9800 (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.

• IHC

Supplier Data

Immunohistochemistry - Anti-Alkaline Phosphatase antibody [EPR27506-72] (BSA and Azide free) (AB305306)

This data was developed using ab305305, the same antibody clone in a different buffer formulation. Immunohistochemical analysis of paraffin-embedded human spleen tissue labeling Alkaline Phosphatase with ab305305 at 1/2000 (0.261 µg/ml) followed by ready to use Leica DS9800 (Bond® Polymer Refine Detection) kit. Low expression in human spleen lymphocytes with positive staining on vascular endothelium. The section was incubated with ab305305 for 30 mins at room temperature. The immunostaining was performed on a Leica Biosystems BOND® RX instrument. Counterstained with Hematoxylin. Secondary antibody only control : Secondary antibody is ready to use Leica DS9800 (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.

• WB

Supplier Data

Western blot - Anti-Alkaline Phosphatase antibody [EPR27506-72] (BSA and Azide free) (AB305306)

This data was developed using ab305305, the same antibody clone in a different buffer formulation. Blocking and diluting buffer and concentration : 5% NFDM/TBST ALPL is a glycoprotein of approximately 80 kDa and detected as a 55 kDa band after treated with Protein Deglycosylation MIX II. Exposure time : 37 seconds

All lanes:

Western blot - Anti-Alkaline Phosphatase antibody [EPR27506-72] (<a href='/en-us/products/primary-antibodies/alkaline-phosphatase-antibody-epr27506-72-ab305305'>ab305305</a>) at 1/1000 dilution

Lane 1:

Untreated human adrenal gland tisuue lysate at 20 µg

Lane 2:

Human adrenal gland tissue lysate treated with Protein Deglycosylation Mix II at 20 µg

Secondary

All lanes:

Goat Anti-Rabbit IgG (HRP) with minimal cross-reactivity with human IgG at 1/2000 dilution

Observed band size: 55 kDa,80 kDa

false

Exposure time: 37s

• WB

Supplier Data

Western blot - Anti-Alkaline Phosphatase antibody [EPR27506-72] (BSA and Azide free) (AB305306)

This data was developed using ab305305, the same antibody clone in a different buffer formulation. Blocking and diluting buffer and concentration : Intercept® (TBS) Blocking Buffer diluted with an equal volume of 0.1% TBS. The samples were run on a Bis-Tris gel. Performed under reducing conditions. False colour image of Western blot : Anti-Alkaline Phosphatase antibody [EPR27506-72] (ab305305) staining at 1/1000 dilution, shown in green; Mouse anti-GAPDH antibody [6C5] (ab8245) loading control staining at 1/20000 dilution, shown in red. In Western blot, ab305305 was shown to bind specifically to Alkaline Phosphatase. A band was observed at 75 kDa in wild-type HAP1 cell lysates whereas no signal observed at this size in Alkaline Phosphatase knockout cell line. To generate this image, wild-type and Alkaline Phosphatase knockout HAP1 cell lysates were analyzed. First, samples were run on a SDS-PAGE gel then transferred onto an immobilon-FL PVDF membrane. Membranes were blocked in in Intercept® (TBS) Blocking Buffer diluted with an equal volume of 0.1% TBS 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) at 1/10000 dilution and Goat anti-Mouse IgG H&L (IRDye® 680RD) preabsorbed (ab216776) at 1/20000 dilution.

All lanes:

Western blot - Anti-Alkaline Phosphatase antibody [EPR27506-72] (<a href='/en-us/products/primary-antibodies/alkaline-phosphatase-antibody-epr27506-72-ab305305'>ab305305</a>) at 1/1000 dilution

Lane 1:

Wild-type HAP1 whole cell lysate at 20 µg

Lane 2:

Alkaline Phosphatase knockout HAP1 whole cell lysate at 20 µg

Lane 3:

Human adrenal gland at 20 µg

Secondary

Lanes 1 - 3:

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 1/10000 dilution

Lanes 1 - 3:

Western blot - Goat anti-Mouse IgG H&L (IRDye® 680RD) preadsorbed (<a href='/en-us/products/secondary-antibodies/goat-mouse-igg-h-l-irdye-680rd-preadsorbed-ab216776'>ab216776</a>) at 1/20000 dilution

Observed band size: 75 kDa

false

• WB

Supplier Data

Western blot - Anti-Alkaline Phosphatase antibody [EPR27506-72] (BSA and Azide free) (AB305306)

This data was developed using ab305305, the same antibody clone in a different buffer formulation. Blocking and diluting buffer and concentration : 5% NFDM/TBST Low expression : A549 (PMID : 33858415 ). Exposure time : Lane 1 : 6 seconds Lanes 2 and 3 : 180 seconds

All lanes:

Western blot - Anti-Alkaline Phosphatase antibody [EPR27506-72] (<a href='/en-us/products/primary-antibodies/alkaline-phosphatase-antibody-epr27506-72-ab305305'>ab305305</a>) at 1/1000 dilution

Lane 1:

Saos-2 whole cell lysate at 20 µg

Lane 2:

HeLa whole cell lysate at 20 µg

Lane 3:

A549 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

Observed band size: 80 kDa

false

Exposure time: 6s