Profiling Human Pluripotent Stem Cell Markers Using Multiplexed Antibody Arrays

ABSTRACT

The Human Pluripotent Stem Cell Antibody Array (Catalog # ARY010) is a rapid and economical tool designed to simultaneously detect the relative levels of 15 different stem cell markers in a single sample. Capture antibodies have been carefully selected for each analyte and spotted in duplicate on nitrocellulose membranes. Cellular extracts are diluted and incubated with the antibody array. The array is washed to remove unbound proteins, followed by incubation with a cocktail of biotinylated detection antibodies. Streptavidin-HRP and chemiluminescent detection reagents are applied, and a signal is produced at each capture spot corresponding to the amount of protein bound. Analysis of undifferentiated and differentiated BG01V cell extracts show changes in stem cell marker levels throughout the differentiation process. The results obtained with the Antibody Array were confirmed by immunocytochemistry for each stage of BG01V differentiation.

Oct-3/4 and GATA-4 expression were also measured using Western blot and RT-PCR for undifferentiated and mesendoderm differentiated BG01V cell extracts. Array experiments can be completed with 5.5 hours of hands-on time and do not require the use of specialized equipment. Thus, the Human Pluripotent Stem Cell Antibody Array offers a sensitive and efficient means to detect changes in multiple protein markers of differentiation that complements other available tools such as PCR and microscopy.

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