|H M R||Endogenous||12||Rabbit|
For western blots, incubate membrane with diluted primary antibody in 5% w/v BSA, 1X TBS, 0.1% Tween® 20 at 4°C with gentle shaking, overnight.
NOTE: Please refer to primary antibody datasheet or product webpage for recommended antibody dilution.
From sample preparation to detection, the reagents you need for your Western Blot are now in one convenient kit: #12957 Western Blotting Application Solutions Kit
NOTE: Prepare solutions with reverse osmosis deionized (RODI) or equivalent grade water.
Load 20 µl onto SDS-PAGE gel (10 cm x 10 cm).
NOTE: Volumes are for 10 cm x 10 cm (100 cm2) of membrane; for different sized membranes, adjust volumes accordingly.
* Avoid repeated exposure to skin.
posted June 2005
revised November 2013
Reprobing of an existing membrane is a convenient means to immunoblot for multiple proteins independently when only a limited amount of sample is available. It should be noted that for the best possible results a fresh blot is always recommended. Reprobing can be a valuable method but with each reprobing of a blot there is potential for increased background signal. Additionally, it is recommended that you verify the removal of the first antibody complex prior to reprobing so that signal attributed to binding of the new antibody is not leftover signal from the first immunoblotting experiment. This can be done by re-exposing the blot to ECL reagents and making sure there is no signal prior to adding the next primary antibody.
NOTE: Prepare solutions with reverse osmosis deionized (RODI) or equivalently purified water.
posted June 2005
revised October 2016
Protocol Id: 10
Supplied in 10 mM sodium HEPES (pH 7.5), 150 mM NaCl, 100 µg/ml BSA and 50% glycerol. Store at –20°C. Do not aliquot the antibody.
MIF Antibody recognizes endogenous levels of total MIF protein.
Human, Mouse, Rat
Polyclonal antibodies are produced by immunizing animals with a synthetic peptide corresponding to residues surrounding Tyr100 of human MIF protein. Antibodies are purified by protein A and peptide affinity chromatography.
MIF (macrophage migration inhibitory factor) is a pleiotropic cytokine that stimulates pro-inflammatory, chemotactic, and growth responses in cells (1). MIF binds its cognate receptor (a CD74/CD44 complex) to activate multiple signaling pathways such as Src, ERK, MAPK, Akt, and suppress p53-induced apoptosis (2). The interaction of MIF with non-cognate chemokine receptors CXCR2 and CXCR4 promotes chemotaxis that enables recruitment of monocytes/neutrophils and T cells (3). During an innate immune response, MIF has been shown to repress the inhibitory effects of glucocorticoids on macrophages and T cells, thus promoting host inflammation and immunity (4). MIF may also play roles in the progression of other disease processes, including cancer cell proliferation and metastasis, and angiogenesis (5), atherosclerotic plaque formation following myocardial ischemia (6), and autoimmune pathogenesis (7). MIF has thus been proposed as a promising therapeutic drug target for multiple indications.
Cell Signaling Technology is a trademark of Cell Signaling Technology, Inc. Tween is a registered trademark of ICI Americas, Inc.
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