Digital quantification using amplified single-molecule detection

Jarvius, Jonas; Melin, Jonas; Göransson, Jenny; Stenberg, Johan A.; Simon, Fabrice; González-Rey, Carlos; Bertilsson, Stefan; Nilsson, Mats

doi:10.1038/nmeth916

Cited by 176 publications

(190 citation statements)

References 15 publications

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“…(39)(40)(41)(42) As mentioned earlier, the 50nt ssDNA aptamer construct employed in this study can be amplified via RCA. Although less sensitive than PCR, the RCA technique is convenient as it can be performed at a constant temperature, e.g.…”

Section: Rca Detectionmentioning

confidence: 93%

Protein detection via direct enzymatic amplification of short DNA aptamers

Fischer

Tarasow

Tok

2008

Analytical Biochemistry

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Aptamers are single-stranded nucleic acids that fold into defined tertiary structures to bind target molecules with high specificities and affinities. DNA aptamers have garnered much interest as recognition elements for biodetection and diagnostic applications due to their small size, ease of discovery and synthesis, and chemical and thermal stability. Herein, we describe the design and application of a short DNA molecule capable of both protein target binding and amplifiable bioreadout processes. As both recognition and readout capabilities are incorporated into a single DNA molecule, tedious conjugation procedures required for protein-DNA hybrids can be omitted. The DNA aptamer is designed to be amplified directly by either the polymerase chain reaction (PCR) or rolling circle amplification (RCA) processes, taking advantage of real-time amplification monitoring techniques for target detection. A combination of both RCA and PCR provides a wide protein target dynamic range (1 μM to 10 pM).

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Section: Rca Detectionmentioning

confidence: 93%

Protein detection via direct enzymatic amplification of short DNA aptamers

Fischer

Tarasow

Tok

2008

Analytical Biochemistry

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“…Moreover, we were able to detect close association of IP 3 R and ANO1 channels using a proximity ligation assay (PLA), a highly specific proteomics method that enables optical detection of closely associated proteins (must be within less than 40 nm proximity in order to be detected; Jarvius et al . 2006; Soderberg et al . 2006).…”

Section: Coupling Between Ano1 and Endoplasmic Reticulum; Is This A Gmentioning

confidence: 99%

Activation of Ca²⁺‐activated Cl⁻ channel ANO1 by localized Ca²⁺ signals

Jin

Shah

et al. 2014

The Journal of Physiology

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Ca2+‐activated chloride channels (CaCCs) regulate numerous physiological processes including epithelial transport, smooth muscle contraction and sensory processing. Anoctamin‐1 (ANO1, TMEM16A) is a principal CaCC subunit in many cell types, yet our understanding of the mechanisms of ANO1 activation and regulation are only beginning to emerge. Ca2+ sensitivity of ANO1 is rather low and at negative membrane potentials the channel requires several micromoles of intracellular Ca2+ for activation. However, global Ca2+ levels in cells rarely reach such levels and, therefore, there must be mechanisms that focus intracellular Ca2+ transients towards the ANO1 channels. Recent findings indeed indicate that ANO1 channels often co‐localize with sources of intracellular Ca2+ signals. Interestingly, it appears that in many cell types ANO1 is particularly tightly coupled to the Ca2+ release sites of the intracellular Ca2+ stores. Such preferential coupling may represent a general mechanism of ANO1 activation in native tissues.

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“…Thus, the development of assay signal amplification paths has become important, and several amplification techniques have been developed, including catalytic nanoparticles (1,2 ), PCR (3,4 ), rolling-circle amplification (5,6 ), enzyme-triggered strand displacement amplification (7)(8)(9)(10), entropydriven signal amplification (11,12 ), and cascade signal amplification (13,14 ). The PCR is currently the most widely used thermal-cycling technique for DNA amplification.…”

confidence: 99%