MA1-029

antibody from Invitrogen Antibodies

Targeting: ACO2 ACONM

Western blot (Supportive data in Antibodypedia)

Immunocytochemistry (Supportive data in Antibodypedia)

Immunohistochemistry (Supportive data in Antibodypedia)

Antibody Data

Product number

MA1-029

Provider

Invitrogen Antibodies

Product name

Aconitase 2 Monoclonal Antibody (7G4)

Antibody type

Monoclonal

Antigen

Recombinant full-length protein

Reactivity

Human, Mouse, Rat

Host

Mouse

Isotype

IgG

Antibody clone number

7G4

Vial size

100 µg

Concentration

1 mg/mL

Storage

-20° C, Avoid Freeze/Thaw Cycles

Western blot

Supportive validation

Submitted by

Invitrogen Antibodies (provider)

Main image

Experimental details

Western blot analysis of Aco-2 was performed by loading 25 µg of various whole cell lysates onto a 4-20% Tris-HCl polyacrylamide gel. Proteins were transferred to a PVDF membrane and blocked with 5% Milk/TBST for at least 1 hour. Membranes were probed with a mouse monoclonal antibody recognizing Aco-2 (Product # MA1-029) at a dilution of 1:1000 overnight at 4°C on a rocking platform. Membranes were washed in TBS-0.1% Tween 20 and probed with a goat anti-mouse-HRP secondary antibody (Product # 31430) at a dilution of 1:20,000 for at least one hour. Membranes were washed and chemiluminescent detection performed using Super Signal West Dura (Product # 34075).

Supportive validation

Submitted by

Invitrogen Antibodies (provider)

Main image

Experimental details

Western blot analysis of Aco-2 was performed by loading 25 µg of whole cell lysate from Aco-2 SMART pool siRNA (Product # L-009566-01-0005) transfected or non-targeting control (Product # D-001810-10) transfected MCF7 cells onto a 4-20% Tris-HCl polyacrylamide gel. Proteins were transferred to a PVDF membrane and blocked with 5% Milk/TBST for at least 1 hour. Membranes were probed with Aco-2 mouse monoclonal antibody (Product # MA1-029) at a dilution of 1:1000 overnight at 4°C on a rocking platform. Membranes were washed in TBS-0.1%Tween 20 and probed with a goat anti-mouse-HRP secondary antibody (Product # 31430) at a dilution of 1:20,000 for at least one hour. Membranes were washed and chemiluminescent detection performed using Super Signal West Dura (Product # 34075).

Immunocytochemistry

Supportive validation

Submitted by

Invitrogen Antibodies (provider)

Main image

Experimental details

Immunofluorescent analysis of ACO2 (green) in 3T3 cells. The cells were fixed with 4% paraformaldehyde for 15 minutes, permeabilized with 0.1% Triton X-100 in PBS for 15 minutes and blocked with 3% Blocker BSA (Product # 37525) in PBS for 30 minutes at room temperature. Cells were stained with or without ACO2 mouse monoclonal antibody (Product # MA1-029), at a concentration of 5 µg/mL for 1 hour at room temperature, and then incubated with a Goat anti-Mouse IgG (H+L) Superclonal Secondary Antibody, Alexa Fluor® 488 conjugate (Product # A28175) at a dilution of 1:1000 for 1 hour at room temperature (both panels, green). Nuclei (both panels, blue) were stained with Hoechst 33342 dye (Product # 62249). Images were taken on a Thermo Scientific ToxInsight at 20X magnification.

Supportive validation

Submitted by

Invitrogen Antibodies (provider)

Main image

Experimental details

Immunofluorescent analysis of Aco-2 using anti-Aco-2 monoclonal antibody (Product # MA1-029) (shown in green) in HeLa cells. Formalin fixed cells were permeabilized with 0.1% Triton X-100 in TBS for 10 minutes at room temperature. Cells were then blocked with 1% Blocker BSA (Product # 37525) for 15 minutes at room temperature. Cells were probed with a mouse monoclonal antibody recognizing Aco-2 (Product # MA1-029), at a dilution of 1:100 for at least 1 hour at room temperature. Cells were then washed with PBS and incubated with DyLight 488 goat-anti-mouse secondary antibody (Product # 35503) at a dilution of 1:400 for 30 minutes at room temperature. Nuclei (blue) were stained with Hoechst 33342 dye (Product # 62249). Images were taken on a Thermo Scientific ArrayScan at 20X magnification.

Immunohistochemistry

Supportive validation

Submitted by

Invitrogen Antibodies (provider)

Main image

Experimental details

Immunohistochemistry was performed on cancer biopsies of deparaffinized human colon, myocardial, hepatic and renal cell carcinoma tissues. To expose target proteins heat induced antigen retrieval was performed using 10mM sodium citrate (pH6.0) buffer for 20 minutes at 95°C. Following antigen retrieval tissues were blocked in 10% normal goat serum (Product # 31873) for 20 minutes at room temperature. Tissues were then probed at a dilution of 1:800 with a mouse monoclonal antibody recognizing Aco-2 (Product # MA1-029) overnight at 4°C in a humidified chamber. Detection was performed using a goat anti-mouse HRP secondary antibody followed by colorimetric detection using DAB substrate. Tissues were counterstained with hematoxylin and prepped for mounting. Images were taken at 400X magnification. Results demonstrate cytoplasmic localization of mitochondrial aconitase 2.