Abstract 6311: An enabling technology to detect low abundance targets in cells and tissues

Abstract

Abstract The rapid advances in sequencing, genomics and bioinformatics have generated a huge amount of genomic data available at our disposal. However, some proteins, the products of those genes, are expressed in cells or tissues at extremely low levels, making the identification and characterization of proteins the bottle neck for biomedical research field. Approximately, 65% of the human proteome is expressed at less than a few thousand copies. Thus, the availability of protein detection probes and tools becomes a critical element to measure lower abundance protein targets in the cells and tissues to identify clinical significance of the particular target proteins. There are several common signal amplification methods available for detecting low abundance protein targets. However, in some cases, their sensitivities are still not satisfactory. Biotin/avidin amplification system has been predominantly used to enhance detection sensitivity but it is still sometimes not sufficient, leading to extremely challenging to accurately measure the low expressed protein targets. There is urgently unmet need to develop a new signal amplification system with enhanced detection sensitivity. We develop the Power Styramide Signal Amplification (PSA) system that can detect extremely low-abundance targets in cells and tissues. The PSA technique is much more sensitive than the traditionally used biotin/avidin and TSA amplification methods. PSA uses a concept that utilizes the catalytic activity of horseradish peroxide (HRP) for covalent deposition of Styramide in situ. The radicals generated from the Styramide based substrates have much higher reactivity towards the target, thus producing a more robust signal and achieve much higher sensitivity towards lower expressed targets. When this technique was compared to directly conjugated antibody, we saw a significant increase in the signal and improved detection limit by 10-100 folds. Another advantage using Styramide-based substrates is that the need of either primary or secondary antibodies gets significantly reduced. You can get robust signal just by using very little antibodies without sacrificing the quality of the fluorescent images. Overall, this new technology enables us to detect low expressed targets in cells or tissues using lower antibody concentration with high detection limit. Citation Format: Deven Patel, Jixiang Liu, Pengfei Dong, Shu Kan, Haiyan Wu, Qin Zhao, Zhenjun Diwu. An enabling technology to detect low abundance targets in cells and tissues [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 6311.