Signal amplification on microarrays: techniques and advances in tyramide signal amplification (TSA)

Karl E. Adler; Mary C. Tyler; Alvydas Mikulskis; Mike O'Malley; Jeff J. Broadbent; Eva E. Golenko; Andy L. Johnson; Steve Lott; Anis H. Khimani; Mark N. Bobrow

doi:10.1117/12.427977

4 June 2001 Signal amplification on microarrays: techniques and advances in tyramide signal amplification (TSA)

Karl E. Adler, Mary C. Tyler, Alvydas Mikulskis, Mike O'Malley, Jeff J. Broadbent, Eva E. Golenko, Andy L. Johnson, Steve Lott, Anis H. Khimani, Mark N. Bobrow

Author Affiliations +

Proceedings Volume 4266, Microarrays: Optical Technologies and Informatics; (2001) https://doi.org/10.1117/12.427977
Event: BiOS 2001 The International Symposium on Biomedical Optics, 2001, San Jose, CA, United States

Abstract

Increased sensitivity for differential mRNA expression analysis on microarrays is rapidly becoming a serious need as the technology matures. Current techniques using direct cyanine labeled targets are effective for expression analysis of abundant mRNA sources but have limited utility for analysis where mRNA quantities are limited. Tyramide signal amplification (TSA^TM) applied to microarray detection provides dramatic improvements in sensitivity, allowing the reduction of sample sizes by as much as 200-fold. The technique includes hapten labeling of two separate RNA populations, microarray hybridization and detection of each hapten with sequential signal amplification steps. The system uses fluorescein and biotin nucleotide analogs as the hapten pair. Hybridized fluorescein and biotin labeled targets are sequentially reacted with horseradish peroxidase and cyanine 3 and cyanine 5 tyramides, resulting in the numerous depositions of these fluorophors on the array. Differential gene expression analysis of LNCaP and PC3 prostate cancer cell lines using one microgram of total RNA and TSA detection, indicates good correlation with results obtained starting with 100 micrograms ((mu) g) of total RNA in a conventional cyanine 3 and cyanine 5 nucleotide analog labeling and detection system (i.e., the direct method).

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Karl E. Adler, Mary C. Tyler, Alvydas Mikulskis, Mike O'Malley, Jeff J. Broadbent, Eva E. Golenko, Andy L. Johnson, Steve Lott, Anis H. Khimani, and Mark N. Bobrow "Signal amplification on microarrays: techniques and advances in tyramide signal amplification (TSA)", Proc. SPIE 4266, Microarrays: Optical Technologies and Informatics, (4 June 2001); https://doi.org/10.1117/12.427977

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KEYWORDS

Analog electronics

Luminescence

Signal detection

Fluorescence spectroscopy

Biological research

Prostate cancer

Statistical analysis

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