MA1-179

antibody from Invitrogen Antibodies

Targeting: EIF5A EIF-5A, EIF5A1, MGC104255, MGC99547

Western blot (Supportive data in Antibodypedia)

Immunocytochemistry (Supportive data in Antibodypedia)

Flow cytometry (Supportive data in Antibodypedia)

Antibody Data

Product number

MA1-179

Provider

Invitrogen Antibodies

Product name

eIF5A Monoclonal Antibody (4E10G8)

Antibody type

Monoclonal

Antigen

Purifed from natural sources

Description

MA1-179 detects EIF5A from human samples.

Antibody clone number

4E10G8

Concentration

1 mg/mL

Western blot

Supportive validation

Submitted by

Invitrogen Antibodies (provider)

Main image

Experimental details

Western blot analysis of EIF5A was performed on whole cell extracts (30 µg lysate) of HEL 92.1.7 (Lane 1), HeLa (Lane 2), NTERA-2 (Lane 3) and THP1 (Lane 4). Protein samples were electrophoresed using a Novex® NuPAGE® 10 % Bis-Tris gel (Product # NP0302BOX), XCell SureLock™ Electrophoresis System (Product # EI0002), and Novex® Sharp Pre-Stained Protein Standard (Product # LC5800). Resolved proteins were then transferred onto a nitrocellulose membrane with the iBlot® 2 Dry Blotting System (Product # IB21001), and blocked with 5% Milk in PBST. The blots were probed with a EIF5A Mouse Monoclonal Antibody (Product # MA1-179, 1-2 µg/mL) followed by a Goat anti-Mouse IgG (H+L) Superclonal™ Secondary Antibody, HRP conjugate (Product # A28177, 0.4 µg/mL, 1:2500 dilution). Chemiluminescent detection was performed using Pierce™ ECL Western Blotting Substrate (Product # 32106). EIF5A was detected at approximately 20kDa across cell lines tested.

Immunocytochemistry

Supportive validation

Submitted by

Invitrogen Antibodies (provider)

Main image

Experimental details

Immunofluorescent analysis of EIF5A was performed using 70% confluent log phase HeLa cells. The cells were fixed with 4% paraformaldehyde for 10 minutes, permeabilized with 0.1% Triton™ X-100 for 10 minutes, and blocked with 1% BSA for 1 hour at room temperature. The cells were then labeled with a EIF5A Mouse Monoclonal Antibody (Product # MA1-179) at 2 µg/mL in 0.1% BSA for 3 hours at room temperature, and then labeled with a Goat anti-Mouse IgG (H+L) Superclonal™ Secondary Antibody, Alexa Fluor® 488 conjugate (Product # A28175) at a dilution of 1:2000 for 45 minutes at room temperature (Panel a: green). Nuclei (Panel b: blue) were stained with SlowFade® Gold Antifade Mountant with DAPI (Product # S36938). F-actin (Panel c: red) was stained with Alexa Fluor® 555 Rhodamine Phalloidin (Product # R415, 1:300). Panel d represents the merged image showing cytoplasmic localization. Panel e shows the no primary antibody control. The images were captured at 60X magnification.

Flow cytometry

Supportive validation

Submitted by

Invitrogen Antibodies (provider)

Main image

Experimental details

Flow cytometry analysis of EIF5A was done on HeLa cells. Cells were fixed with 70% ethanol for 10 minutes, permeabilized with 0.25% Triton™ X-100 for 20 minutes, and blocked with 5% BSA for 30 minutes at room temperature. Cells were then labeled with a EIF5A Mouse Monoclonal Antibody (Product # MA1-179, red histogram) or a mouse isotype control (pink histogram) at 3-5 µg/million cells in 2.5% BSA. After incubation at room temperature for 2 hours, the cells were then labeled with a Rabbit anti-Mouse IgG (H+L) Secondary Antibody, Alexa Fluor® 488 conjugate (Product # A-11059) at a dilution of 1:400 for 30 minutes at room temperature. The representative 10,000 cells were acquired and analyzed for each sample using an Attune® Acoustic Focusing Cytometer. The purple histogram represents unstained control cells and the green histogram represents no primary antibody control.