Revvity, AL3210C, AlphaLISA Human and Mouse LC3B-II Detection Kit, 500 assay points

Overview
LC3 is a mammalian homolog of the yeast ATG8 protein and is distributed throughout mammalian tissues and cultured cells. LC3 is a key component of the autophagy process - a degradative program that maintains cellular homeostasis. During autophagy, cytoplasmic components such as cytosolic proteins and organelles are engulfed by autophagosomes Concurrently, a cytosolic form of LC3 (LC3-I) is conjugated to phosphatidylethanolamine to form LC3-phosphatidylethanolamine conjugate (LC3-II). This newly formed conjugate is recruited to autophagosomal membranes. The subsequent fusion of autophagosomes with lysosomes results in the formation of autolysosomes, and intra-autophagosomal components are degraded by lysosomal hydrolases. At the same time, LC3-II within the autolysosomal lumen is degraded. The LC3-II level has been found to be proportional to autophagosome abundance making LC3-II the most popular marker of autophagy.
Formats:

Our 100 assay point kit allows you to run 100 wells in 96-well format, using a 100 µL reaction volume (10 µL of sample).
Our 500 assay point kit allows you to run 500 wells in 96-well or 384-well format, using a 50 µL reaction volume (5 µL of sample).
Our 5,000 assay point kit allows you to run 5,000 wells in 96-well or 384-well format, using a 50 µL reaction volume (5 µL of sample).

Features:

No-wash steps, no separation steps
ELISA alternative technology
Sensitive detection
Broad sample compatibility
Small sample volume

AlphaLISA technology allows the detection of molecules of interest in a no-wash, highly sensitive, quantitative assay. In an AlphaLISA assay, a biotinylated anti-analyte antibody binds to the Streptavidin-coated Donor beads while another anti-analyte antibody is conjugated to AlphaLISA Acceptor beads. In the presence of the analyte, the beads come into close proximity. The excitation of the Donor beads causes the release of singlet oxygen molecules that triggers a cascade of energy transfer in the Acceptor beads, resulting in a sharp peak of light emission at 615 nm.
How it works
Principle of the AlphaLISA human and mouse LC3B-II detection kit
The AlphaLISA Human and Mouse LC3B-II assay is based on an AlphaLISA sandwich immunoassay. A biotinylated anti-LC3B-II antibody binds to the streptavidin coated AlphaLISA Donor beads, while an anti- LC3B-II antibody is conjugated to AlphaLISA Acceptor beads.
In the presence of LC3B-II, the beads come into proximity. The excitation of the Donor beads provokes the release of singlet oxygen molecules that triggers a cascade of energy transfer within the Acceptor beads, resulting in emission with λmax at 615 nm.

 

Protocol of the AlphaLISA human and mouse LC3B-II assay
The AlphaLISA Human and Mouse LC3B-II assay can be run in a 96- or 384-well detection plate (50 µL final). As described here, samples (cell lysates or tissue lysates) or control lysate are dispensed directly into the assay plate for the detection of LC3B-II by AlphaLISA reagents. No washing steps are needed. The protocol can be further miniaturized or upscaled by simply resizing each addition volume proportionally.
Application-Protein Quantification
Automation Compatible-Yes
Brand-AlphaLISA
Detection Modality-Alpha
Protocol Time-Overnight at RT
Sample Volume-5 µL
Shipping Conditions-Shipped in Blue Ice
Target-LC3B-II
Target Class-Biomarkers
Target Species-Human Mouse
Technology-Alpha
Unit Size-500 assay points