MA1-20881

antibody from Invitrogen Antibodies

Targeting: MIF GIF, GLIF

Western blot (Supportive data in Antibodypedia)

Immunocytochemistry (Supportive data in Antibodypedia)

Antibody Data

Product number

MA1-20881

Provider

Invitrogen Antibodies

Product name

MIF Monoclonal Antibody (2Ar3)

Antibody type

Monoclonal

Antigen

Recombinant full-length protein

Description

Store product as a concentrated solution. Centrifuge briefly prior to opening the vial.

Reactivity

Human

Host

Mouse

Isotype

IgG

Antibody clone number

2Ar3

Vial size

100 µg

Concentration

0.5 mg/mL

Storage

Store at 4°C short term. For long term storage, store at -20°C, avoiding freeze/thaw cycles.

Western blot

Supportive validation

Submitted by

Invitrogen Antibodies (provider)

Main image

Experimental details

Western blot was performed using Anti-MIF Monoclonal Antibody (2Ar3) (Product # MA1-20881) and a 15 kDa band corresponding to Macrophage migration inhibitory factor was observed across cell lines tested and was upregulated in U-87 MG upon VEGF treatment. Whole cell extracts (30 µg lysate) of THP-1 (Lane 1), U-937 (Lane 2), U-87 MG (Lane 3), U-87 MG treated with VEGF (100 ng/mL for 48hrs) (Lane 4), Jurkat (Lane 5), HEK-293 (Lane 6) were electrophoresed using NuPAGE™ 12% Bis-Tris Protein Gel (Product # NP0342BOX). 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 µg/mL) and detected by chemiluminescence with Goat anti-Mouse IgG (H+L) Superclonal™ Recombinant Secondary Antibody, HRP (Product # A28177, 1:6000 dilution) using the iBright FL 1000 (Product # A32752). Chemiluminescent detection was performed using Novex® ECL Chemiluminescent Substrate Reagent Kit (Product # WP20005).

Immunocytochemistry

Supportive validation

Submitted by

Invitrogen Antibodies (provider)

Main image

Experimental details

Immunofluorescence analysis of Macrophage migration inhibitory factor was performed using 70% confluent log phase U-87 MG 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 45 minutes at room temperature. The cells were labeled with MIF Monoclonal Antibody (2Ar3) (Product # MA1-20881) at 1:100 dilution in 0.1% BSA, incubated at 4 degree celsius overnight and then labeled with Donkey anti-Mouse IgG (H+L) Highly Cross-Adsorbed Secondary Antibody, Alexa Fluor Plus 488 (Product # A32766), (1:2000 dilution), 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, 1:300). Panel d represents the merged image showing cytoplasmic and secreted localization. Panel e represents control cells with no primary antibody to assess background. The images were captured at 60X magnification.