Anti-Insulin antibody [RM1019]

• IHC-P

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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Insulin antibody [RM1019] (AB282459)

Immunohistochemical analysis of paraffin-embedded Human skeletal muscle tissue labeling Insulin with ab282459 at 1/10000 (0.055 ug/ml) followed by a ready to use Goat Anti-Rabbit IgG H&L (HRP polymer). The section was incubated with ab282459 overnight at 4°C. Counterstained with Hematoxylin.

Negative control : no staining in human skeletal muscle.

Secondary antibody only control : Secondary antibody is a ready to use Goat Anti-Rabbit IgG H&L (HRP polymer).

Heat mediated antigen retrieval using ab93678 (citrate buffer, pH 6.0).

• IHC-P

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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Insulin antibody [RM1019] (AB282459)

Immunohistochemical analysis of paraffin-embedded Human pancreas tissue labelling Insulin with ab282459 at 1/10000 (0.055 ug/ml) followed by a ready to use Goat Anti-Rabbit IgG H&L (HRP polymer). Positive staining on the islets in human pancreas. The section was incubated with ab282459 overnight at 4°C. Counterstained with Hematoxylin.

Secondary antibody only control : Secondary antibody is a ready to use Goat Anti-Rabbit IgG H&L (HRP polymer).

Heat mediated antigen retrieval using ab93678 (citrate buffer, pH 6.0).

• IHC-Fr

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Immunohistochemistry (Frozen sections) - Anti-Insulin antibody [RM1019] (AB282459)

Immunohistochemical analysis of 4% PFA-fixed, 0.2% Triton X-100 permeabilized frozen Mouse pancreas tissue labeling Insulin with ab282459 at 1/1000 (0.565 ug/ml) dilution followed by ab150077 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) at 1/1000 (2 ug/ml) dilution (Green). Cytoplasmic staining on mouse pancreas islet is observed. The nuclear counterstain was DAPI (Blue).

Secondary antibody control : Secondary antibody is ab150077 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) at 1/1000 (2 ug/ml) dilution.

Heat mediated antigen retrieval using sodium citrate buffer (10mM citrate pH 6.0 + 0.05% Tween-20).

• ICC/IF

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Immunocytochemistry/ Immunofluorescence - Anti-Insulin antibody [RM1019] (AB282459)

Immunofluorescent analysis of 4% Paraformaldehyde-fixed, 0.1% TritonX-100 permeabilized Beta-TC-6 cells labelling Insulin with ab282459 at 1/1000 (0.551 ug/ml) dilution, followed by ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed antibody at 1/1000 (2 ug/ml) dilution (Green). Confocal image showing cytoplasmic staining in Beta-TC-6 cells. ab195889 Anti-alpha Tubulin mouse monoclonal antibody - Microtubule Marker (Alexa Fluor® 594) was used to counterstain tubulin at 1/200 (2.5 ug/ml) dilution (Red). The Nuclear counterstain was DAPI (Blue).

Negative control : C2C12 (PMID : 22973301)

Secondary antibody only control : Secondary antibody is ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed at 1/1000 (2 ug/ml) dilution.

• IHC-P

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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Insulin antibody [RM1019] (AB282459)

Immunohistochemical analysis of paraffin-embedded Mouse pancreas tissue labeling Insulin with ab282459 at 1/10000 (0.055 ug/ml) followed by a ready to use Goat Anti-Rabbit IgG H&L (HRP polymer). Positive staining on the islet in mouse pancreas. The section was incubated with ab282459 overnight at 4°C. Counterstained with Hematoxylin.

Secondary antibody only control : Secondary antibody is a ready to use Goat Anti-Rabbit IgG H&L (HRP polymer).

Heat mediated antigen retrieval using ab93678 (citrate buffer, pH 6.0).

• IHC-Fr

Supplier Data

Immunohistochemistry (Frozen sections) - Anti-Insulin antibody [RM1019] (AB282459)

Immunohistochemical analysis of 4% PFA-fixed, 0.2% Triton X-100 permeabilized frozen Rat pancreas tissue labeling Insulin with ab282459 at 1/1000 (0.551 ug/ml) dilution followed by ab150077 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) at 1/1000 (2 ug/ml) dilution (Green). Cytoplasmic staining on rat pancreas islet is observed. The nuclear counterstain was DAPI (Blue).

Secondary antibody control : Secondary antibody is ab150077 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488)at 1/1000 (2 ug/ml) dilution.

Heat mediated antigen retrieval using sodium citrate buffer (10mM citrate pH 6.0 + 0.05% Tween-20).

• Flow Cyt (Intra)

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Flow Cytometry (Intracellular) - Anti-Insulin antibody [RM1019] (AB282459)

Flow cytometric analysis of 4% paraformaldehyde fixed 90% methanol permeabilized C2C12 (Mouse myoblasts myoblast) (Left) / Beta-TC-6 (Mouse pancreas insulinoma beta cell) (Right) cells labelling Insulin with ab282459 at 1/5000 dilution (0.1ug) (Red) compared with a Rabbit monoclonal IgG (ab172730) (Black) isotype control and an unlabelled control (cells without incubation with primary antibody and secondary antibody) (Blue). A Goat F(ab')2 Anti-Rabbit IgG(DyLight® 488, ab98507) at 1/500 dilution was used as the secondary antibody.

Negative control : C2C12 (PMID : 22973301)

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Insulin antibody [RM1019] (AB282459)

Immunohistochemical analysis of paraffin-embedded Rat pancreas tissue labeling Insulin with ab282459 at 1/10000 (0.055 ug/ml) followed by a ready to use Goat Anti-Rabbit IgG H&L (HRP polymer). Positive staining on the islet in rat pancreas. The section was incubated with ab282459 overnight at 4°C. Counterstained with Hematoxylin.

Secondary antibody only control : Secondary antibody is a ready to use Goat Anti-Rabbit IgG H&L (HRP polymer).

Heat mediated antigen retrieval using ab93678 (citrate buffer, pH 6.0).

• IP

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Immunoprecipitation - Anti-Insulin antibody [RM1019] (AB282459)

Insulin was immunoprecipitated from 0.35 mg Mouse pancreas lysate 10 ug with ab282459 at 1/30 dilution (2ug in 0.35mg lysates). Western blot was performed on the immunoprecipitate using ab282459 at 1/1000 dilution. VeriBlot for IP secondary antibody(HRP)(ab131366) was used at 1/5000 dilution.

Lane 1 : Mouse pancreas lysate 10 ug

Lane 2 : ab282459 IP in Mouse pancreas lysate

Lane 3 : Rabbit monoclonal IgG (ab172730) instead of ab282459 in Mouse pancreas lysate

Blocking and dilution buffer and concentration : 5% NFDM/TBST.

Exposure time : 3 min.

All lanes:

Immunoprecipitation - Anti-Insulin antibody [RM1019] (ab282459)

Predicted band size: 12 kDa

Observed band size: 12 kDa

false

• WB

Lab

Western blot - Anti-Insulin antibody [RM1019] (AB282459)

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

Negative control : C2C12 (PMID : 22973301)

Exposure time : 26 seconds.

All lanes:

Western blot - Anti-Insulin antibody [RM1019] (ab282459) at 1/1000 dilution

Lane 1:

Beta-TC-6 (Mouse pancreas insulinoma beta cell) whole cell lysate at 20 µg

Lane 2:

C2C12 (Mouse myoblasts myoblast) whole cell lysate at 20 µg

Secondary

All lanes:

Goat Anti-Rabbit IgG, (H+L), Peroxidase conjugated (<a href='/ko/products/secondary-antibodies/goat-rabbit-igg-h-l-hrp-ab97051'>ab97051</a>) at 1/100000 dilution

Predicted band size: 12 kDa

Observed band size: 12 kDa

false

• I-ELISA

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Indirect ELISA - Anti-Insulin antibody [RM1019] (AB282459)

ELISA using ab282459 at varying antibody concentrations and antigen concentration at 1000 ng/ml. An Alkaline Phosphatase-conjugated AffiniPure Goat Anti-Rabbit IgG (H+L) at 1/2500 was used as the secondary antibody.

• WB

Lab

Western blot - Anti-Insulin antibody [RM1019] (AB282459)

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

This blot was developed using a higher sensitivity ECL substrate.

Exposure time : 3 minutes.

All lanes:

Western blot - Anti-Insulin antibody [RM1019] (ab282459) at 1/1000 dilution

Lane 1:

Human pancreas lysate at 20 µg

Lane 2:

Mouse pancreas lysate at 20 µg

Lane 3:

Rat pancreas lysate at 20 µg

Secondary

All lanes:

Goat Anti-Rabbit IgG, (H+L), Peroxidase conjugated (<a href='/ko/products/secondary-antibodies/goat-rabbit-igg-h-l-hrp-ab97051'>ab97051</a>) at 1/100000 dilution

Predicted band size: 12 kDa

Observed band size: 12 kDa

false