Analyzing the phosphorylation profile of kinases and their protein substrates is essential to understand how cells recognize and respond to changes in their environment. The NSB PhosphoKinase Microarray is a rapid, sensitive, and economical tool to unveil the relative site-specific phosphorylation of the major kinases without performing labor intensive and multiple immunoprecipitations and Western blots. Twenty two capture antibodies were judiciously selected, and each capture antibody is spotted in four replicates per chamber area. There are two chamber areas on the standard (1" x 3") slide (up and down half), and the format is compatible with most chambers including Agilent 2 microarray hybridization chamber. The level of the phosphorylated protein is assessed by using the phospho-specific antibodies and the fluorescence detection.
NSB PhosphoKinase Microarray contains a major 22 kinds of anti-phosphokinase antibodies printed on an NSB27 slide, NSB's proprietary NanoCone surface. Due to the desirable regular spacing between the immobilized molecules, antibodies printed on it ensures an enhanced interaction with a target molecule and thus provides with higher sensitivity and reliability.
Typically 30~50 μg of total cell extract is used for the detection of specific phoshokinases in the NSB microarray. Superb performance includes use of small sample amount of the total extract from cell line sample, tissue sample, and primary cell culture sample and the low variation of the array signal in four replicate spots (C.V. less than 10%, mostly <5%).
NSB PhosphoKinase microarray guarantees the good spot morphology and enhance stability (stable up to 6 months when stored at -20”). Nevertheless, it is recommended to use within a few months. When stored at 4”, usage in one month would produce the best result.
Figure 1. Selected antibodies and the printing format of NSB PhosphoKinase Microarray.
Figure 2. A typical step for NSB PhosphoKinase Microarray.
Figure 3. A result of NSB PhosphoKinase Microarray with a cell line extract.
Figure 4. A result of NSB PhosphoKinase Microarray with a tissue extract.
Figure 5. A result of NSB PhosphoKinase Microarray with a primary cell culture extract.
Use of the NSB27 NanoCone coated slide.
Judicious selection of 22 phospho-specific antibodies.
Microarray format for the high throughput expression profiling.
Four antibody replicates per chamber area, two chamber areas on the standard slide (up and down half).
Good for a protein extract from cell, tissue, or serum.
High specificity and sensitivity as well as low C.V. of the fluorescence signal.
Compatible with all standard fluorescent microarray scanners.
An Excel template file for C.V. analysis is provided.
Obtain protein expression profiling (Relative)
Measure changes in phosphorylation status at a specific site
Compare profiles of normal, diseased or treated samples
Identify candidate biomarkers