Compaction of rolling circle amplification products increases signal integrity and signal–to–noise ratio

• Rolling circle amplification (RCA) for generation of distinct fluorescent signals in situ relies upon the self-collapsing properties of single-stranded DNA in commonly used RCA-based methods. By introducing a cross-hybridizing DNA oligonucleotide during rolling circle amplification, we demonstrate that the fluorophore-labeled RCA products (RCPs) become smaller. The reduced size of RCPs increases the local concentration of fluorophores and as a result, the signal intensity increases together with the signal-to-noise ratio. Furthermore, we have found that RCPs sometimes tend to disintegrate and may be recorded as several RCPs, a trait that is prevented with our cross-hybridizing DNA oligonucleotide. These effects generated by compaction of RCPs improve accuracy of visual as well as automated in situ analysis for RCA based methods, such as proximity ligation assays (PLA) and padlock probes.

Subject headings

TEKNIK OCH TEKNOLOGIER  -- Medicinteknik -- Medicinsk bildbehandling (hsv//swe)

ENGINEERING AND TECHNOLOGY  -- Medical Engineering -- Medical Image Processing (hsv//eng)

NATURVETENSKAP  -- Biologi -- Annan biologi (hsv//swe)

NATURAL SCIENCES  -- Biological Sciences -- Other Biological Topics (hsv//eng)

NATURVETENSKAP  -- Biologi (hsv//swe)

NATURAL SCIENCES  -- Biological Sciences (hsv//eng)

MEDICIN OCH HÄLSOVETENSKAP  -- Medicinsk bioteknologi (hsv//swe)

MEDICAL AND HEALTH SCIENCES  -- Medical Biotechnology (hsv//eng)

Keyword

Datoriserad bildbehandling

Computerized Image Processing

Publication and Content Type

ref (subject category)

art (subject category)