- MA5-29070 - Invitrogen Antibodies CA2 antibody

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Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Western blot analysis of Carbonic Anhydrase II in Lane A: 293T Whole Cell Lysate (30 µg). Samples were probed using a Carbonic Anhydrase II Monoclonal Antibody (Product # MA5-29070) at a 1:10000 dilution, followed by a Goat Anti-Rabbit IgG (H+L), Dylight 800 Secondary Antibody at a 1:10000 dilution. Western blot was performed under reducing conditions. Predicted band size:29 kDa. Observed band size:30 kDa.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Western Blot using Carbonic Anhydrase II Recombinant Rabbit Monoclonal Antibody (1) (Product # MA5-29070) at 1:10,000 dilution. Lane A: 293T Whole Cell Lysate. Lysates/proteins at 30 μg per lane. Secondary Goat Anti-Rabbit IgG H&L (DyLight™ 800) at 1:10,000 dilution. Developed using the Odyssey technique. Performed under reducing conditions. Predicted band size: 29 kDa. Observed band size: 30 kDa.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Western blot was performed using Anti-Carbonic Anhydrase II Monoclonal Antibody (Product # MA5-29070) and ~30kDa band corresponding to CA2 was observed across cell lines tested except in SW480 and T-47D along with an uncharacterized band (*) at ~60 kDa. Whole cell extracts (30 µg lysate) of HEK-293 (Lane 1), THP-1 (Lane 2), A-431 (Lane 3), SW480 (Lane 4) and T-47D (Lane 5) were electrophoresed using NuPAGE® 10% Bis-Tris gel (Product # NP0302BOX). Resolved proteins were then transferred onto a nitrocellulose membrane (Product # IB23001) by iBlot® 2 Dry Blotting System (Product # IB21001).The blot was probed with the primary antibody (1:10000 dilution) and detected by chemiluminescence with Goat anti-Rabbit IgG (H+L), Superclonal™ Recombinant Secondary Antibody, HRP (Product # A27036, 1:4000 dilution) using the iBright FL 1000 (Product # A32752). Chemiluminescent detection was performed using Novex® ECL Chemiluminescent Substrate Reagent Kit (Product # WP20005).

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Immunofluorescence analysis of CA2 was performed using 70% confluent log phase HEK-293 cells. The cells were fixed with 4% paraformaldehyde for 10 minutes, permeabilized with 0.1% Triton™ X-100 for 15 minutes, and blocked with 2% BSA for 1 hour at room temperature. The cells were labeled with Carbonic Anhydrase II Recombinant Rabbit Monoclonal Antibody (Product # MA5-29070) at 1:100 dilution in 0.1% BSA, incubated at 4 degree Celsius overnight and then labeled with Goat anti-Rabbit IgG (H+L) Superclonal™ Recombinant Secondary Antibody, Alexa Fluor® 488 conjugate (Product # A27034) at a dilution of 1:2000 for 45 minutes at room temperature (Panel a: green). Nuclei (Panel b: blue) were stained with ProLong™ Diamond Antifade Mountant with DAPI (Product # P36962). F-actin (Panel c: red) was stained with Rhodamine Phalloidin (Product # R415). Panel d represents the merged image showing Cytoplasmic localization. Panel e represents control cells with no primary antibody to assess background. The images were captured at 60X magnification.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Carbonic Anhydrase II Immunoprecipitation using: Lane A: 0.5 mg 293T Whole Cell Lysate 2 µL with Carbonic Anhydrase II Recombinant Rabbit Monoclonal Antibody (1) (Product # MA5-29070) and 15 µL of 50 % Protein G agarose. Primary antibody: Carbonic Anhydrase II Recombinant Rabbit Monoclonal Antibody (1), at 1:2,000 dilution. Secondary antibody: Dylight 800-labeled antibody to rabbit IgG (H+L), at 1:5,000 dilution. Developed using the Odyssey technique. Performed under reducing conditions. Predicted band size: 29 kDa. Observed band size: 29 kDa.

MA5-29070