Anti-Amylin/DAP antibody [EPR22556-138]

• IHC-P

Unknown

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Amylin/DAP antibody [EPR22556-138] (AB254259)

Immunohistochemical analysis of paraffin-embedded human pancreas tissue labeling Amylin/DAP with ab254259 at 1/2000 dilution, followed by ready to use Rabbit specific IHC polymer detection kit HRP/DAB (ab209101). Cytoplasmic staining on the islet in human pancreas (PMID : 30014749). The section was incubated with ab254259 for 30 mins at room temperature. The immunostaining was performed on a Leica Biosystems BOND^® RX instrument. Counterstained with hematoxylin.

Secondary antibody only control : Used PBS instead of primary antibody, secondary antibody is ready to use Rabbit specific IHC polymer detection kit HRP/DAB (ab209101).

Heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0, epitope retrieval solution 2) for 20 mins.

• mIHC

Lab

Multiplex immunohistochemistry - Anti-Amylin/DAP antibody [EPR22556-138] (AB254259)

Multiplex immunohistochemistry analysis of formalin/PFA-fixed paraffin-embedded rat pancreas tissue staining Trypsin with ab325233 at a 1/500 (0.98 μg/ml) dilution, ab317606 anti-SHANK2 used at a 1/100 (5.21 μg/ml) dilution and ab254259 anti-Amylin used at a 1/2000 (0.26 μg/ml) dilution.

Panel A : anti-Trypsin (green; Opal™690), anti-SHANK2 (magenta; Opal™520), anti-Amylin (gray; Opal™570) on rat pancreas.

Panel B : anti-Trypsin staining exocrine gland in rat pancreas.

Panel C : anti-SHANK2 staining pancreatic ducts in rat pancreas.

Panel D : anti-Amylin staining islet cells in rat pancreas.

Nuclear DNA was labeled with DAPI (shown in blue).

The section was incubated in three rounds of staining : in the order of ab325233, ab317606 and ab254259 for 30 mins at room temperature. Each round was followed by a separate fluorescent tyramide signal amplification system.

The immunostaining was performed on a Leica Biosystems BOND® RX instrument with an Opal™ 4-color kit. Image acquisition was performed with Leica SP8 confocal microscope.

Heat mediated antigen retrieval was performed with Tris-EDTA buffer (pH 9.0, Epitope Retrieval Solution2) for 20 mins.

• mIHC

Lab

Multiplex immunohistochemistry - Anti-Amylin/DAP antibody [EPR22556-138] (AB254259)

Multiplex immunohistochemistry analysis of formalin/PFA-fixed paraffin-embedded rat pancreas tissue staining Robo4 with ab300046 at a 1/100 (3.75 μg/ml) dilution, ab325233 anti-Trypsin used at a 1/500 (0.98 μg/ml) dilution and ab254259 anti-Amylin used at a 1/2000 (0.26 μg/ml) dilution.

Panel A : anti-Robo4 (green; Opal™570), anti-Trypsin (magenta; Opal™690), anti-Amylin (gray; Opal™570) on rat pancreas.

Panel B : anti-Robo4 staining endothelium in rat pancreas.

Panel C : anti-Trypsin staining exocrine gland in rat pancreas.

Panel D : anti-Amylin staining islet cells in rat pancreas.

Nuclear DNA was labeled with DAPI (shown in blue).

The section was incubated in three rounds of staining : in the order of ab300046, ab325233 and ab254259 for 30 mins at room temperature. Each round was followed by a separate fluorescent tyramide signal amplification system.

The immunostaining was performed on a Leica Biosystems BOND® RX instrument with an Opal™ 4-color kit. Image acquisition was performed with Leica SP8 confocal microscope.

Heat mediated antigen retrieval was performed with Tris-EDTA buffer (pH 9.0, Epitope Retrieval Solution2) for 20 mins.

• ICC/IF

Unknown

Immunocytochemistry/ Immunofluorescence - Anti-Amylin/DAP antibody [EPR22556-138] (AB254259)

Immunofluorescent analysis of 4% paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized Beta-TC-6 (mouse pancreas insulinoma beta cell) cells labeling Amylin/DAP with ab254259 at 1/1000 dilution, followed by Goat Anti-Rabbit IgG H&L (Alexa Fluor^® 488) (ab150077) secondary antibody at 1/1000 dilution (green). Confocal image showing cytoplasmic and membranous staining in Beta-TC-6 cell line. Negative control : NIH/3T3 (PMID : 15802374). The nuclear counter stain is DAPI (blue). Counterstained with ab195889 Anti-alpha Tubulin antibody [DM1A] - Microtubule Marker (Alexa Fluor^® 594) at a 1/200 dilution (red). The negative control is the secondary antibody only.

• mIHC

Lab

Multiplex immunohistochemistry - Anti-Amylin/DAP antibody [EPR22556-138] (AB254259)

Multiplex immunohistochemistry analysis of formalin/PFA-fixed paraffin-embedded mouse pancreas tissue staining Robo4 with ab300046 at a 1/100 (3.75 μg/ml) dilution, ab325233 anti-Trypsin used at a 1/500 (0.98 μg/ml) dilution and ab254259 anti-Amylin used at a 1/2000 (0.26 μg/ml) dilution.

Panel A : anti-Robo4 (green; Opal™570), anti-Trypsin (magenta; Opal™690), anti-Amylin (gray; Opal™570) on mouse pancreas.

Panel B : anti-Robo4 staining endothelium in mouse pancreas.

Panel C : anti-Trypsin staining exocrine gland in mouse pancreas.

Panel D : anti-Amylin staining islet cells in mouse pancreas.

Nuclear DNA was labeled with DAPI (shown in blue).

The section was incubated in three rounds of staining : in the order of ab300046, ab325233 and ab254259 for 30 mins at room temperature. Each round was followed by a separate fluorescent tyramide signal amplification system.

The immunostaining was performed on a Leica Biosystems BOND® RX instrument with an Opal™ 4-color kit. Image acquisition was performed with Leica SP8 confocal microscope.

Heat mediated antigen retrieval was performed with Tris-EDTA buffer (pH 9.0, Epitope Retrieval Solution2) for 20 mins.

• IHC-P

Unknown

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Amylin/DAP antibody [EPR22556-138] (AB254259)

Immunohistochemical analysis of paraffin-embedded rat pancreas tissue labeling Amylin/DAP with ab254259 at 1/2000 dilution, followed by ready to use Rabbit specific IHC polymer detection kit HRP/DAB (ab209101). Cytoplasmic staining on the islet in rat pancreas (PMID : 30014749). The section was incubated with ab254259 for 30 mins at room temperature. The immunostaining was performed on a Leica Biosystems BOND^® RX instrument. Counterstained with hematoxylin.

Secondary antibody only control : Used PBS instead of primary antibody, secondary antibody is ready to use Rabbit specific IHC polymer detection kit HRP/DAB (ab209101).

Heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0, epitope retrieval solution 2) for 20 mins.

• IHC-Fr

Unknown

Immunohistochemistry (Frozen sections) - Anti-Amylin/DAP antibody [EPR22556-138] (AB254259)

Immunohistochemical analysis of 4% PFA-fixed, 0.2% Triton X-100 permeabilized frozen mouse pancreas tissue labeling Amylin/DAP with ab254259 at 1/500 dilution (green), followed by ab150077 AlexaFluor^®488 Goat anti-Rabbit secondary at a 1/1,000 dilution. Positive staining on beta cells of mouse pancreatic islets (PMID : 14656003) is observed. Counterstained with DAPI (blue). Secondary antibody only control : Used PBS instead of primary antibody, secondary antibody is ab150077 AlexaFluor^®488 Goat anti-Rabbit used at a 1/1,000 dilution. Heat mediated antigen retrieval using sodium citrate buffer (10 mM citrate pH 6.0 and 0.05% Tween-20).

• IHC-P

Unknown

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Amylin/DAP antibody [EPR22556-138] (AB254259)

Immunohistochemical analysis of paraffin-embedded mouse pancreas tissue labeling Amylin/DAP with ab254259 at 1/2000 dilution, followed by ready to use Rabbit specific IHC polymer detection kit HRP/DAB (ab209101). Cytoplasmic staining on the islet in mouse pancreas (PMID : 30014749). The section was incubated with ab254259 for 10 mins at room temperature. The immunostaining was performed on a Leica Biosystems BOND^® RX instrument. Counterstained with hematoxylin.

Secondary antibody only control : Used PBS instead of primary antibody, secondary antibody is ready to use Rabbit specific IHC polymer detection kit HRP/DAB (ab209101).

Heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0, epitope retrieval solution 2) for 20 mins.

• IHC-Fr

Unknown

Immunohistochemistry (Frozen sections) - Anti-Amylin/DAP antibody [EPR22556-138] (AB254259)

Immunohistochemical analysis of 4% PFA-fixed, 0.2% Triton X-100 permeabilized frozen rat pancreas tissue labeling Amylin/DAP with ab254259 at 1/500 dilution (green), followed by ab150077 AlexaFluor^®488 Goat anti-Rabbit secondary at a 1/1,000 dilution. Positive staining on beta cells of rat pancreatic islets (PMID : 14656003) is observed. Counterstained with DAPI (blue). Secondary antibody only control : Used PBS instead of primary antibody, secondary antibody is ab150077 AlexaFluor^®488 Goat anti-Rabbit used at a 1/1,000 dilution. Heat mediated antigen retrieval using sodium citrate buffer (10 mM citrate pH 6.0 and 0.05% Tween-20).

• Flow Cyt (Intra)

Unknown

Flow Cytometry (Intracellular) - Anti-Amylin/DAP antibody [EPR22556-138] (AB254259)

Intracellular flow cytometric analysis of 4% paraformaldehyde-fixed, 90% methanol permeabilized NIH/3T3 (Mouse embryonic fibroblast, Left) / Beta-TC-6 (Mouse pancreas insulinoma beta cell, Right) cell lines labeling Amylin/DAP with ab254259 at 1/500 (red) compared with a Rabbit monoclonal IgG isitype control (ab172730) (black) and an unlabeled control (cells without incubation with primary antibody and secondary antibody) (blue). Goat anti rabbit IgG (Alexa Fluor^® 488, ab150077), at 1/2000 dilution was used as the secondary antibody. Negative control : NIH/3T3 (PMID : 15802374).

• IP

Unknown

Immunoprecipitation - Anti-Amylin/DAP antibody [EPR22556-138] (AB254259)

Amylin/DAP was immunoprecipitated from 0.35 mg Beta-TC-6 (mouse pancreas insulinoma beta cell) whole cell lysate with ab254259 at 1/30 dilution. Western blot was performed from the immunoprecipitate using ab254259 at 1/1000 dilution. VeriBlot for IP Detection Reagent (HRP) (ab131366), was used at 1/5000 dilution.

Lane 1 : Beta-TC-6 whole cell lysate 10 μg (Input).
Lane 2 : ab254259 IP in Beta-TC-6 whole cell lysate.
Lane 3 : Rabbit monoclonal IgG (ab172730) instead of ab254259 in Beta-TC-6 whole cell lysate.

Blocking/Dilution buffer : 5% NFDM/TBST.
Exposure time : 3 minutes.

All lanes:

Immunoprecipitation - Anti-Amylin/DAP antibody [EPR22556-138] (ab254259)

Predicted band size: 10 kDa

Observed band size: 10 kDa

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