1. LUMIER-Assays Dr. Manfred Koegl Protein Interaction Unit
Genomics And Proteomics Core Facility

2. LUMIER assays
are a reliable, cheap, high-throughput method to measure protein-protein interactions in mammalian cells.
The assay tests the interaction of two proteins which are transiently expressed in mammalian cells. Like classical co-immuneprecipitations, the method is based on a co-purification protocol.
Idea by Barrios-Rodiles et al., (2005) Science 307:

3. LUMIER-Assays
Two tagged proteins (here labelled X and Y), are transiently expressed in mammalian cells.
Cells are lysed, and the first tag is used for affinity purification of protein X on magnetic beads.
The second tag is a luciferase and used for detection of protein Y.
IgG-coated
magnetic
beads Y
Luci
affinity tag X
*”LUMIER”´, Idea by Barrios-Rodiles et al., (2005) Science 307:

4. LUMIER workflow Expression constructs for
1. Transiently transfect into HEK293 cells
Expression constructs for
protein Y (luciferase-tagged) + / - protein X (protein A-tagged)
2. Prepare cell lysate, split in 2 aliquots
3. Allow binding to magnetic beads
4. Wash
5. Measure luciferase activity
Calculate bound luciferase relative to total luciferase

5. Controls
sample Y
negative control X Y
(hexuplicates +/- s.d.)