Western blot analysis of extracts from various cell lines and tissues using Lysozyme C-1/2 Antibody (upper) and α-Actinin (D6F6) XP® Rabbit mAb #6487 (lower).
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.
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
Lysozyme C-1/2 Antibody recognizes endogenous levels of total mouse and rat Lysozyme C-1 and Lysozyme C-2 proteins.Species Reactivity:
Polyclonal antibodies are produced by immunizing animals with a synthetic peptide corresponding to residues surrounding Ala60 of mouse Lysozyme C-1/2 protein. Antibodies are purified by peptide affinity chromatography.
Lysozymes are secreted proteins that have bacteriolytic function which are critical for mammalian innate immune function. All lysozymes function to defend host animals from microbial infection by hydrolyzing bacterial cell wall peptidoglycan (1). Conventional-type lysozymes (Lysozyme C) are one of three types of lysozymes; each family member is categorized based on amino acid sequence and biochemical properties. Lysozyme C is expressed in mammalian secretions like tears, urine, and milk, but are also expressed by phagocytes such as macrophages, neutrophils, and dendritic cells. Lysozyme C is encoded in humans by a single LYZ gene. The mouse orthologs of Lysozyme C are encoded by two genes, Lyz1 and Lyz2, which encode Lysozyme C-1 and Lysozyme C-2 (Lysozyme C-1/2). Interestingly, Lyz2 is upregulated in microglia of Alzheimer's disease mouse model brains that have been stimulated by specific forms of activity (2). Lyz1 and Lyz2 are uniquely expressed in microglia, and increased Lyz2 correlates with microglia-mediated β-amyloid (Aβ) clearance, suggesting that Lysozyme C-1/2 may directly contribute to microglial-clearance of Aβ or act as a marker for certain microglial activity states in the brain (3).
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