Confocal immunofluorescent analysis of HeLa cells, chloroquine-treated (50 μM, overnight; left), nutrient-starved with EBSS (4 hr, middle), or untreated (right) using LC3A/B (D3U4C) XP® Rabbit mAb (Alexa Fluor® 555 Conjugate) #13173 (red) and β-Actin (8H10D10) Mouse mAb #3700 (green). Blue pseudocolor = DRAQ5® #4084 (fluorescent DNA dye).
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This Cell Signaling Technology antibody is conjugated to Alexa Fluor® 555 fluorescent dye and tested in-house for direct immunofluorescent analysis in human cells. The antibody is expected to exhibit the same species cross-reactivity as the unconjugated LC3A/B (D3U4C) XP® Rabbit mAb #12741.
Supplied in PBS (pH 7.2), less than 0.1% sodium azide and 2 mg/ml BSA. Store at 4°C. Do not aliquot the antibody. Protect from light. Do not freeze.
NOTE: Prepare solutions with reverse osmosis deionized (RODI) or equivalently purified water.
NOTE: Cells should be grown, treated, fixed and stained directly in multi-well plates, chamber slides or on coverslips.
NOTE: All subsequent incubations should be carried out at room temperature unless otherwise noted in a humid light-tight box or covered dish/plate to prevent drying and fluorochrome fading.
posted December 2010
Protocol Id: 221
LC3A/B (D3U4C) XP® Rabbit mAb (Alexa Fluor® 555 Conjugate) recognizes endogenous levels of total LC3A and LC3B proteins.
Human, Mouse, Rat
Monkey, Xenopus, Bovine, Dog, Pig
Monoclonal antibody is produced by immunizing animals with a synthetic peptide corresponding to residues surrounding Leu44 of human LC3B protein (conserved in LC3A).
Autophagy is a catabolic process for the autophagosomic-lysosomal degradation of bulk cytoplasmic contents (1,2). Autophagy is generally activated by conditions of nutrient deprivation, but it has also been associated with a number of physiological processes including development, differentiation, neurodegenerative diseases, infection, and cancer (3). Autophagy marker Light Chain 3 (LC3) was originally identified as a subunit of microtubule-associated proteins 1A and 1B (termed MAP1LC3) (4) and subsequently found to contain similarity to the yeast protein Apg8/Aut7/Cvt5 critical for autophagy (5). Three human LC3 isoforms (LC3A, LC3B, and LC3C) undergo post-translational modifications during autophagy (6-9). Cleavage of LC3 at the carboxy terminus immediately following synthesis yields the cytosolic LC3-I form. During autophagy, LC3-I is converted to LC3-II through lipidation by a ubiquitin-like system involving Atg7 and Atg3 that allows for LC3 to become associated with autophagic vesicles (6-10). The presence of LC3 in autophagosomes and the conversion of LC3 to the lower migrating form, LC3-II, have been used as indicators of autophagy (11).
Explore pathways + proteins related to this product.
Cell Signaling Technology is a trademark of Cell Signaling Technology, Inc.
XP is a registered trademark of Cell Signaling Technology, Inc.
Alexa Fluor is a registered trademark of Life Technologies Corporation.
The Alexa Fluor dye antibody conjugates in this product are sold under license from Life Technologies Corporation for research use only, except for use in combination with DNA microarrays. The Alexa Fluor® dyes (except for Alexa Fluor® 430 dye) are covered by pending and issued patents. Alexa Fluor® is a registered trademark of Molecular Probes, Inc.