Two-Color Labeling
and Fluorescent Detection Kit
Two-Color Labeling
and Fluorescent Detection
The Whatman Two-Color Labeling and Fluorescent Detection Kit is designed to label two protein samples. The labeled proteins are pooled and probed against arrayed antibodies in a competitive binding assay and detected using indirect fluorescence.
The kit is intended for use with the 2 pad FAST Slides, including the Serum Biomarker Chip. The kit contains the Universal Linkage System (ULS) chemistry to label samples containing approximately 250 µg of protein in serum, plasma or a whole cell lysate. The kit is designed to label two different protein samples, each with a different hapten. Sufficient labeling reagent is provided to perform a hapten swapping experiment.
Features
- Highly efficient and uniform labeling of complex serum samples
- Reproducible labeling and signal detection
- Stable, robust, and fast non enzymatic procedure
- Reduces pH dependency of labeling efficiency
- System solution includes labeling reagents, fluorescent conjugate, and bench-friendly protocol
- Accounts for hapten-specific differences in either Biotin-ULS or Fluorescein-ULS labeling efficiencies
- Averages differences in antibody-antigen binding interactions caused by steric hindrance
- Minimizes chip-to-chip variability – includes an internal control within the assay
The first pad on the slide is probed with a mixture of two different protein samples, each labeled with a different hapten; the second pad is probed with the same two protein samples but with the haptens reversed. The normalized intensity for each element of each pad is calculated as the average of the biotin and fluorescein labeled derived intensities from a two pad experiment. The ratio between the signal intensity at each spot corresponds to the concentration ratio of the proteins found in the two samples. This method is attractive for antibody chips as it takes into account any hapten-specific differences in antigen-antibody interactions.
The use of the ULS labeling system minimizes background by using indirect fluorescence detection, labels multiple amino acids, and requires no additional materials or reagents.
Each kit includes 20 μL Biotin-ULS, 20 μl Fluorescein-ULS, 80 μl 10x Protein Labeling Buffer, 80 μl 10x KREAstop Solution, 150 μl Streptavidin-DY 647 Conjugate, 350 μl Anti-Fluorescein Antibody-DY 547 Conjugate, 8 Micro Bio-Spin Chromatography Columns, and a User's Manual.